Differential acute effects of aldosterone, dexamethasone, and hyperkalemia on distal tubular potassium secretion in the rat kidney.

To determine the specific effects on renal potassium transport of acute elevations in plasma aldosterone, dexamethasone, and potassium concentrations, we studied adrenalectomized rats prepared such that each factor could be varied independently. Clearance data alone could not be used to deduce the underlying tubular transport effects, however, since infusion of each of these agents was associated with a marked change in urinary flow rate, which may itself have influenced potassium excretion. We therefore used a technique of continuous microperfusion, in vivo, of single superficial distal tubules to evaluate potassium secretion at constant luminal flow rate during each experimental maneuver. Acute aldosterone infusion was associated with a 90% stimulation of potassium secretion by microperfused tubules. However, total kidney sodium excretion and urinary flow rate were markedly reduced, and these factors opposed the direct tubular action of aldosterone, resulting in no net change in the amount of potassium excreted into the final urine. Conversely, dexamethasone had no direct effect on potassium secretion by single microperfused tubules, but it caused a sharp increase in urinary flow and sodium excretion, and secondarily enhanced urinary potassium excretion by 50%. Hyperkalemia per se stimulated renal potassium excretion both via a direct tubular effect and by increasing urinary flow rate. We conclude that urinary potassium excretion after infusion of each of these agents represents the net result of direct tubular effects and secondary flow-mediated changes.

Quantification of the rat spinal microglial response to peripheral nerve injury as revealed by immunohistochemical image analysis and flow cytometry.

Microgliosis is implicated in the pathophysiology of several neurological disorders, including neuropathic pain. Consequently, perturbation of microgliosis is a mechanistic and drug development target in neuropathic pain, which highlights the requirement for specific, sensitive and reproducible methods of microgliosis measurement. In this study, we used the spinal microgliosis associated with L5 spinal nerve transection and minocycline-induced attenuation thereof to: (1) evaluate novel software based semi-quantitative image analysis paradigms for the assessment of immunohistochemical images. Microgliosis was revealed by immunoreactivity to OX42. Several image analysis paradigms were assessed and compared to a previously validated subjective categorical rating scale. This comparison revealed that grey scale measurement of the proportion of a defined area of spinal cord occupied by OX42 immunoreactive cells is a robust image analysis paradigm. (2) Develop and validate a flow cytometric approach for quantification of spinal microgliosis. The flow cytometric technique reliably quantified microgliosis in spinal cord cell suspensions, using OX42 and ED9 immunoreactivity to identify microglia. The results suggest that image analysis of immunohistochemical revelation of microgliosis reliably detects the spinal microgliosis in response to peripheral nerve injury and pharmacological attenuation thereof. In addition, flow cytometry provides an alternative approach for quantitative analysis of spinal microgliosis elicited by nerve injury.

Simulations of electron transfer in the NADPH-bound catalase from Proteus mirabilis PR.

Catalase-bound NADPH both prevents and reverses the accumulation of compound II, an inactive form of catalase that is generated from the normal active intermediate form (compound I) when catalase is exposed to a steady flow of hydrogen peroxide. The mechanism for the regeneration reaction is unknown although NADPH could act either as a one-electron or a two-electron donor. Recently, a reaction scheme has been proposed in which the formation of compound II from compound I generates a neighboring radical species within the protein. NADPH would then donate two electrons, one to compound II for reduction of the iron and the other to the protein free radical. In this paper, we report calculations to find the dominant electron tunneling pathways between NADPH and the heme iron in the catalase from the peroxide-resistant mutant of Proteus mirabilis. Two major tunneling pathways are found which fuse together on Ser-196. It is suggested that the sequence Gly-Ser of the loop that divides the beta 5-strand is the key element for shielding a radical amino acid.

Effect of immunodepletion of MHC class II-positive cells from pancreatic islets on generation of cytotoxic T-lymphocytes in mixed islet-lymphocyte coculture.

In vitro manipulation of pancreatic islets to decrease islet immunogenicity before transplantation has largely been directed at eliminating the major histocompatibility complex (MHC) class II-positive passenger leukocytes from the islets. The mixed islet-lymphocyte coculture (MILC) system was used to quantitate the efficacy of immunodepletion of MHC class II-positive cells from pancreatic islets in terms of reducing immunogenicity. With these experiments we compared the in vitro immunogenicity of MHC class II-depleted islets with untreated islets. B10.BR (H-2k) islets were treated with anti-Iak alloserum followed by complement. This treatment successfully eliminated MHC class II-positive cells from the islets, as demonstrated by indirect immunofluorescence techniques. Depleted islets generated slightly lower amounts of allospecific cytotoxic T-lymphocyte (CTL) activity when exposed to C57BL/6 (H-2b) splenocytes in the MILC than untreated control islets. Although the amount of CTL generated by the depleted islets was slightly less than that generated by untreated islets, there was significant stimulation of CTL by the MHC class II-depleted islets. Therefore, the presence or absence of MHC class II cells within the islet is unlikely to be the decisive factor contributing to islet immunogenicity.

Effect of 15-deoxyspergualin on immediate function and long-term survival of transplanted islets in murine recipients of a marginal islet mass.

15-Deoxyspergualin (DSG), a macrophage immunomodulatory agent, was used as a probe in a murine model of islet transplantation to examine 1) the significance of the nonspecific, macrophage-mediated effector arm of beta-cell injury in recipients of a marginal mass of isologous islets by analyzing the duration of temporary posttransplant hyperglycemia, a parameter of immediate beta-cell function; and 2) whether long-term (> 100 days) functional survival could be achieved in recipients of a marginal mass of allogeneic islets. A dose-response study of the number of islets required to ameliorate diabetes showed that 150 isologous islets per recipient resulted in a 75% incidence of cure at a mean of 39.2 +/- 5.8 days posttransplant. DSG-treated (0.625 mg.kg-1.day-1 intraperitoneally) recipients of isologous islets demonstrated a significant (P < 0.01) reduction in the duration of temporary posttransplant hyperglycemia (16.8 +/- 3.2 vs. 39.2 +/- 5.8 days), and DSG-treated recipients of allogeneic islets demonstrated a significant (P < 0.03) improvement in the rate of achieving long-term functional survival (75 vs. 22% in untreated control animals). Finally, identical rates of islet engraftment were found among control animals and DSG-treated animals by measurement of tissue insulin content in transplanted specimens. The results are consistent with the hypothesis that DSG alters the duration of temporary posttransplant hyperglycemia and extends long-term functional survival in murine recipients of a marginal mass of islets, not by affecting the efficiency of islet engraftment, but by suppression of the inhibitory effects on beta-cell function by nonspecific, macrophage mediators.

Analysis of the low-frequency normal modes of the R state of aspartate transcarbamylase and a comparison with the T state modes.

Aspartate transcarbamylase (ATCase) is a classic example of an allosteric enzyme. It catalyzes the conversion of aspartate to carbamyl aspartate, which is the first substrate in the biosynthesis of pyrimidines. Although ATCase is well characterized, both structurally and biochemically, little is known at the atomic level about the large amplitude motions that govern its T-->R quaternary transition. We present the results of calculations of the very-low-frequency normal modes of the CTP-ligated R state ATCase, and we compare them with the equivalent modes in the CTP-ligated T state ATCase. The large-amplitude, delocalized modes of frequencies below 4 cm-1 contribute a large fraction of the atomic fluctuations observed experimentally. They show some ability to drive the R-state structure towards the T-state structure, by promoting some of the quaternary structure rearrangements that take place during the allosteric process. Their potential role in the T-->R transition is quantified and compared with the role of the low-frequency modes of the T state in the quaternary rearrangement.

Analysis of the low frequency normal modes of the T-state of aspartate transcarbamylase.

Aspartate transcarbamylase (ATCase) is an important control enzyme in the pyrimidine biosynthetic pathway in Escherichia coli. It is a classic example of an allosteric protein and has been extensively studied biochemically, kinetically and structurally. As yet, however, a detailed model for the cooperative transition between the tensed (T) and relaxed (R) forms of the protein does not exist. In this work we have calculated the low frequency normal modes of the CTP-ligated T-state of ATCase with the aim of identifying some of the motions that could be important in initiating the transition. The calculated modes, of frequencies lower than 5 per cm, produce root-mean-square coordinate deviations for the atoms which are a substantial fraction of those derived from the crystallographic B-factors. Some of the modes result in displacements which change the quaternary structure of the protein (in particular the elongation of the protein and the relative rotation of the subunits) in such a way that the R-state structure is approached. The implication of these mode motions for the overall T-->R transition process is discussed.

Measuring quality of care at the end of life.

Caring for patients at the end of life presents a series of quality-of-care problems to the health care system. In the past, concern has focused on overaggressive treatment of dying patients. Given rapid changes in the financing and delivery of care, it is time to focus on a range of quality problems and address ways to improve care and achieve outcomes desired by patients and their families. We provide a framework for conceptualizing such a task. This article addresses the purposes of measurement, definition of the patient population, timing of measurement, use of surrogates in measurement, scope of services to be evaluated, and the choice of measures. It emphasizes the necessary links between quality measurement and quality improvement.

Ligand diffusion in the catalase from Proteus mirabilis: a molecular dynamics study.

The role of the channels and cavities present in the catalase from Proteus mirabilis (PMC) was investigated using molecular dynamics (MD) simulations. The reactant and products of the reaction, H(2)O(2) -->1/2 O(2) + H(2)O, catalyzed by the enzyme were allowed to diffuse to and from the active site. Dynamic fluctuations in the structure are found necessary for the opening of the major channel, identified in the X-ray model, which allows access to the active site. This channel is the only pathway to the active site observed during the dynamics, and both the products and reactant use it. H(2)O and O(2) are also detected in a cavity defined by the heme and Ser196, which could play an important role during the reaction. Free energy profiles of the ligands diffusing through the major channel indicate that the barriers to ligand diffusion are less than 20 kJ mol(-1) for each of the species. It is not clear from our study that minor channels play a role for access to the protein active site or to the protein surface.

Direct comparison of cognitive facilitation by physostigmine and tetrahydroaminoacridine in two primate models.

Cognitive facilitation by physostigmine and tetrahydroaminoacridine (THA) was compared in two primate models. Disruption of spatial delayed response performance by scopolamine (0.03 mg/kg) was fully reversed by coadministration of 5 doses of physostigmine in the range 0.03-0.08 mg/kg, but by only one dose (4.0 mg/kg) of THA; partial reversal of some effects of scopolamine was observed at 1 and 3 mg/kg of THA. Visual recognition memory was enhanced following treatment with 4 doses of physostigmine in the range 0.001-0.03 mg/kg. The effect of THA across the group of animals was not significant but performance tended to improve using a dose of 0.8 mg/kg. Our findings indicate that THA does not have a superior profile to physostigmine as a cognitive enhancer in primates.

Gabapentin and pregabalin, but not morphine and amitriptyline, block both static and dynamic components of mechanical allodynia induced by streptozocin in the rat.

A single injection of streptozocin (50 mg/kg, i.p.) led to the development of static and dynamic allodynia in the rat. The two responses were detected, respectively, by application of pressure using von Frey hairs or lightly stroking the hind paw with a cotton bud. Static allodynia was present in the majority of the animals within 10 days following streptozocin. In contrast, dynamic allodynia took almost twice as long to develop and was only present in approximately 60% of rats. Morphine (1-3 mg/kg, s.c.) and amitriptyline (0.25-2.0 mg/kg, p.o.) dose-dependently blocked static allodynia. However, neither of the compounds was effective against dynamic allodynia. In contrast, gabapentin (10-100 mg/kg, p.o.) and the related compound pregabalin (3-30 mg/kg, p.o.) dose-dependently blocked both types of allodynia. However, the corresponding R-enantiomer (10-100 mg/kg, p.o.) of pregabalin, was found to be inactive. The intrathecal administration of gabapentin dose-dependently (1-100 microg/animal) blocked both static and dynamic allodynia. In contrast, administration of similar doses of gabapentin into the hind paw failed to block these responses. It is suggested that in this model of neuropathic pain dynamic allodynia is mediated by A beta-fibres and the static type involves small diameter nociceptive fibres. These data suggest that gabapentin and pregabalin possess a superior antiallodynic profile than morphine and amitriptyline, and may represent a novel class of therapeutic agents for the treatment of neuropathic pain.

Detection of static and dynamic components of mechanical allodynia in rat models of neuropathic pain: are they signalled by distinct primary sensory neurones?

In the present study, chronic constrictive injury (CCI model) of the sciatic nerve or tight ligation of L5 and L6 spinal nerves (Chung model) produced both dynamic and static components of mechanical allodynia in rats. The two responses were detected, respectively, by lightly stroking the hind paw with cotton wool or application of pressure using von Frey hairs. Animals with spinal nerve ligation developed both types of responses at a faster rate compared to animals with the CCI. Morphine (1-3 mg/kg, s.c.) dose-dependently blocked static but not dynamic allodynia. In contrast, pregabalin (previously S-isobutylgaba and CI-1008) dose-dependently (3-30 mg/kg, p.o.) blocked both types of allodynia. In CCI animals, two administrations of capsaicin (100 microg/50 microl) into the plantar surface of the ipsilateral paw at 1-h intervals blocked the maintenance of thermal hyperalgesia without affecting either static or dynamic allodynia. The similar administration of a further two doses of capsaicin into the same animals blocked the maintenance of static allodynia without affecting the dynamic response. These data indicate that thermal hyperalgesia, static and dynamic allodynia are respectively signalled by C-, Adelta- and Abeta/capsaicin insensitive Adelta- primary sensory neurones. It is suggested that pregabalin possesses a superior antiallodynic profile than morphine and may represent a novel class of therapeutic agents for the treatment of neuropathic pain.

Enadoline, a selective kappa-opioid receptor agonist shows potent antihyperalgesic and antiallodynic actions in a rat model of surgical pain.

Enadoline is a highly selective and potent kappa-opioid receptor agonist. This report describes and compares the activities of enadoline and morphine in a rat model of postoperative pain. A 1 cm incision through the muscle and skin of the plantar surface of the right hind paw induced thermal hyperalgesia as well as static and dynamic allodynia lasting at least 2 days. Postoperative testing was carried out using the plantar test for thermal hyperalgesia, von Frey hairs for static allodynia and light stroking with a cotton bud for dynamic allodynia. A single i.v. dose of enadoline 15 min before surgery dose-dependently (1-100 microg/kg) blocked the development of thermal hyperalgesia as well as static and dynamic allodynia for over 24 h with respective MEDs of < or = 1, 10 and 10 microg/kg. The administration of enadoline (100 microg/kg, i.v.), 1 h after surgery, completely blocked the maintenance of the hyperalgesic and allodynic responses, but its duration of action was much shorter (2 h) than when administered before surgery. Previous studies have shown that administration of morphine (1-6 mg/kg, s.c.) 0.5 h before surgery can prevent the development of thermal hyperalgesia with a MED of < or =1 mg/kg, but it has little effect on static allodynia. In the present study similar administration of morphine (1-3 mg/kg), unlike enadoline, had no effect on the development of dynamic allodynia. Morphine dose-dependently (1-6 mg/kg, s.c.) potentiated isoflurane-induced sleeping time and respiratory depression in the rat. However, whilst enadoline also (1-1000 microg/kg, i.v.) potentiated isoflurane-induced sleeping time, it did not cause respiratory depression. It is suggested that enadoline may possess therapeutic potential as a pre-emptive antihyperalgesic and antiallodynic agent.

Utilization of an intramolecular hydrogen bond to increase the CNS penetration of an NK(1) receptor antagonist.

This paper describes the synthesis and physical and biological effects of introducing different substituents at the alpha-position of the tryptophan containing neurokinin-1 receptor antagonist [(R)-2-(1H-indol-3-yl)-1-methyl-1-((S)-1-phenyl-ethylcarbamoyl)-ethyl]-carbamic acid benzofuran-2-ylmethyl ester (CI 1021). The described compounds all exhibit less than 5 nM binding affinities for the human neurokinin-1 receptor and selectivity over the tachykinin NK(2) and NK(3) receptor subtypes. Application of variable temperature nuclear magnetic resonance spectroscopy studies of the amide and urethane protons was utilized to determine the existence of an intramolecular hydrogen bond. This intramolecular hydrogen bond increases the apparent lipophilicity to allow increased central nervous system penetration and pharmacological activity (gerbil foot tap test) in the case of the highest affinity compound [(S)-1-dimethylaminomethyl-2-(1H-indol-3-yl)-1-((S)-1-phenyl-ethylcarbamoyl)-ethyl]-carbamic acid benzofuran-2-ylmethyl ester (PD 174424) over those analogues that could not form an intramolecular hydrogen bond.

[(Aminomethyl)aryloxy]acetic acid esters. A new class of high-ceiling diuretics. 4. Substituted 6,7-dichloro-2,3-dihydrobenzofurans derived by ring annelation.

Ring annelation of the [(aminomethyl)aryloxy]acetic acids produced a series of substituted 6,7-dichloro-2,3-dihydrobenzofuran-2-carboxylic acids. Pharmacologic evaluation of these compounds in rats and dogs indicated that several congeners are extremely potent salidiuretics. Clearance and micropuncture experiments in rats for compound 5a confirmed the high-ceiling diuretic profile and demonstrated that 5a has a site of action at the thick ascending limb of Henle's loop.

Second generation "peptoid" CCK-B receptor antagonists: identification and development of N-(adamantyloxycarbonyl)-alpha-methyl-(R)-tryptophan derivative (CI-1015) with an improved pharmacokinetic profile.

We have previously described the design and development of CI-988, a peptoid analogue of CCK-4 with excellent binding affinity and selectivity for the CCK-B receptor. Due to its anxiolytic profile in animal models of anxiety, this compound was developed as a clinical candidate. However, during its development, it was determined that CI-988 had low bioavailability in both rodent and nonrodent species. In the clinic, it was further established that CI-988 had poor bioavailability. Thus, there was a need to identify an analogue with an improved pharmacokinetic (PK) profile. The poor bioavailability was attributed to poor absorption and efficient hepatic extraction. We envisaged that reducing the molecular weight of the parent compound (5, MW = 614) would lead to better absorption. Thus, we synthesized a series of analogues in which the key alpha-methyltryptophan and adamantyloxycarbonyl moieties, required for receptor binding, were kept intact and the C-terminus was extensively modified. This SAR study led to the identification of tricyclo[3.3.1.1(3,7)]dec-2-yl [1S-[1 alpha(S*)2 beta]-[2-[(2-hydroxycyclohexyl)amino]-1-(1H-indol-3- ylmethyl)-1-methyl-2-oxoethyl]carbamate (CI-1015, 31) with binding affinities of 3.0 and 2900 nM for the CCK-B and CCK-A receptors, respectively. The compound showed CCK-B antagonist profile in the rat ventromedial hypothalamus assay with a Ke of 34 nM. It also showed an anxiolytic like profile orally in a standard anxiety paradigm (X-maze) with a minimum effective dose (MED) of 0.1 microgram/kg. Although the compound is less water soluble than CI-988, oral bioavailability in rat was improved nearly 10 times relative to CI-988 when dosed in HP beta CD. The blood-brain permeability of CI-1015 (31) was also enhanced relative to CI-988 (5). On the basis of the overall improved pharmacokinetic profile as well as enhanced brain penetration, CI-1015 (31) was chosen as a development candidate.

Cold-storage preservation of the canine and rat pancreas prior to islet isolation.

Cold-storage preservation of the canine pancreas prior to islet isolation has previously been noted to reduce the intrasplenic islet autograft success rate; but the mechanism of this deleterious effect has not been determined. We undertook a study in both outbred dogs and Lewis (RT1-1) rats to determine the influence of cold-storage preservation interval, preservation solution, and flushing technique on islet yield and islet viability. The preservation solutions used were those that had proved most efficacious in preserving segmental canine pancreases--namely, the modifications of silica gel fractionated plasma (SGF-III and SGF-IV) and an hydroxyethylstarch/lactobionate solution (UW-1). In the first set of experiments, the traditional vascular flush was used; this was followed by storage at 4 degrees C. After brief periods of preservation (3 hr in the rat, 12 hr in the dog) there was a significant (P less than 0.006) reduction in islet yield. The reduced yields were similar with each solution tested, were made worse with increasing intervals of storage, and resulted in a significant reduction in autograft success rate. The second set of experiments examined the effect of using an intraductal flush prior to preservation, along with the effect of adding collagenase to the preservation fluid. Islet yields were maintained at control values in both animal models using preservation intervals of up to 24 hr. These islet yields produced auto- or isograft success rates similar to those obtained by transplanting freshly obtained tissue; verifying adequate islet viability. We recommend that a pre-storage ductal flush technique be used for cold-storage preservation of the pancreas prior to islet isolation and transplantation.

Differential effects of preexisting uremia and a synchronous kidney graft on pancreas allograft functional survival in rats.

Pancreas transplant results have been better in uremic recipients of a simultaneous kidney than in nonuremic recipients of a pancreas alone. We studied the relative effect of uremia versus a double transplant on functional survival by performing bladder-drained pancreas transplants alone (PTA), kidney transplants alone (KTA), and simultaneous pancreas/kidney (SPK) transplants from Buffalo donors to diabetic Lewis rat recipients that were or were not made uremic 2-3 weeks before by 1 4/5 native nephrectomy. Pancreas graft exocrine function was monitored by urinary amylase (UA). In the PTA and SPK recipients made diabetic by streptozotocin, endocrine function was monitored by measuring nonfasting plasma glucose (PG) levels. Kidney graft function was monitored by plasma creatinine (Cr). Rejection of the endocrine pancreas was defined as an increase of PG to greater than 200 mg/dl; of the exocrine pancreas, as a decline in UA to less than 6000 U/L or to less than 100 U/24 hr; and of the kidney, as an elevation of Cr to greater than 3 mg/dl. The mean functional survival times (MST) of both the endocrine (12.0 +/- 2.1 versus 10.1 +/- 1.1 days, P = 0.036) and exocrine (8.0 +/- 2.1 versus 6.3 +/- 1.3 days, P = 0.016) components of the pancreas grafts were significantly longer in SPK than in PTA recipients. The MST of kidney allografts, however, was not significantly longer in nonuremic SPK than nonuremic KTA recipients (6.7 +/- 1.4 versus 5.7 +/- 0.7 days, P = 0.13). In parallel experiments in recipients immunosuppressed with cyclosporine, the graft survival times were longer, but the relative differences between the PTA, SPK, and KTA groups persisted. Histologically, lymphocyte infiltration began in the two organs almost simultaneously, but the severity of the rejection was more vigorous in the kidney than in the pancreas in doubly grafted rats, and destruction of pancreas grafts progressed more slowly in SPK than in PTA recipients. Preexisting uremia delayed pancreas rejection in both SPK (exocrine 10.6 +/- 2.3, P = 0.032, and endocrine 14.8 +/- 3.4 days, P = 0.065, versus nonuremics) and PTA (exocrine 8.5 +/- 1.7, P = 0.007, and endocrine 12.6 +/- 2.5, P = 0.026, versus nonuremics) nonimmunosuppressed recipients. The MST of kidney grafts was not significantly longer in uremic (8.9 +/- 2.8 days) than in nonuremic (6.7 +/- 1.4 days) SPK recipients (P = 0.081). A synchronous kidney transplant and uremia independently down-modulate the rejection response to a pancreas graft, and a simultaneous pancreas graft has no detrimental effect on the survival of a kidney graft.(ABSTRACT TRUNCATED AT 400 WORDS)

Purified canine islet autografts. Functional outcome as influenced by islet number and implantation site.

Using a modification of the basic principles of pancreatic intraductal collagenase digestion and density gradient purification to isolate canine islets, in conjunction with simultaneous fluorogenic and dithizone islet staining, we quantified the yield, purity, and viability of the isolated islets. We then determined the combined influences of total and weight-corrected islet counts and implantation site on immediate and long-term functional outcome of purified canine islet autografts. Weight-corrected islet counts were 100% sensitive and specific in differentiating successful and unsuccessful islet autografts implanted to the liver (n = 10) and spleen (n = 10) of pancreatectomized dogs. The threshold number of islets required to achieve normoglycemia in the liver (4400 islets/kg) and spleen (4650 islets/kg) were nearly identical. Islet autografts failed to ameliorate hyperglycemia when implanted to the renal subcapsular space (n = 5) at counts of 4400 to 5500 islets/kg. The mean one- and three-month intravenous glucose tolerance test K-values of dogs with purified islet autografts to the liver (-1.43 +/- 0.27 and -1.69 +/- 0.27, respectively) and spleen (-1.78 +/- 0.36 and -1.64 +/- 0.3, respectively) were also similar. Time needed to achieve normoglycemia , however, was significantly (P less than 0.02) shorter for intrahepatic islets (1.0 +/- 0.0 days posttransplant) than intrasplenic islets (6.8 +/- 2.3 days posttransplant). The long-term durability of islet autograft function was not unlimited. Overall, thirteen canine islet autograft recipients have been followed for greater than or equal to 12 months posttransplant (range 12-18 months), seven canine islet autograft recipients (five intrahepatic and two intrasplenic) have had spontaneous recurrence of hyperglycemia at 2, 6, 11, 13, 14, 8, and 16 months, respectively. The phenomenon depended only on the number of islets implanted. The data underscore the significance of quantitatively defined islet preparations and the importance of islet number and implantation site on immediate and long-term functional outcome of canine islet autografts.

Beneficial and detrimental effects of RBC-adsorbed antilymphocyte globulin and prednisone on purified canine islet autograft and allograft function.

We have examined the effects of prednisone, cyclosporine, azathioprine, and RBC-adsorbed goat antidog antilymphocyte globulin on islet graft function in totally pancreatectomized canines with purified, quantitatively defined, autologous, or allogeneic islets transplanted to the liver. The objectives were twofold: (1) to determine the potential detrimental effects to islet autograft function of the aforementioned agents, and (2) to determine the relative efficacy of the "nontoxic" agents in prolonging purified islet allograft function administered in doses that would be considered tolerable in human. The islet autograft studies demonstrated that prednisone given in doses of 1-2 mg/kg/day had a detrimental effect on islet autograft function, and that the combinations of immunosuppression involving CsA, azathioprine, and ALG were not detrimental to islet autograft function to the extent that hyperglycemia would ensue. In the subsequent allograft studies, three groups of canines received islet transplants: (1) controls (n = 5; 7860 +/- 750 islets/kg/weight), (2) canines given CsA and azathioprine (n = 6; 6810 +/- 890 islets/kg/body weight), and (3) canines given CsA, azathioprine, and RBC-adsorbed goat antidog ALG (n = 8; 6540 +/- 710 islets/kg/body weight). The mean (+/- SE) day of rejection (serum glucose greater than or equal to 200 mg/dl) in the group of canine islet allograft recipients receiving CsA, azathioprine, and ALG was 11.8 +/- 1.4 days--significantly prolonged versus islet allograft recipients receiving no immunosuppression (mean survival 4.8 +/- 1.1 days, P less than 0.03), and versus allograft recipients receiving CsA/azathioprine without ALG (mean survival 4.4 +/- 1.4 days, P less than 0.05). Prednisone appears to be detrimental to islet graft function, even at low doses. ALG was not toxic, and significantly extended the survival of canine islet allografts. The inclusion of steroids as part of maintenance immunosuppression, or as treatment for acute rejection of islets, in human islet transplants should be reconsidered, whereas ALG or other antilymphocyte agents should continue to be used.