Early barriers to neonatal porcine islet engraftment in a dual transplant model.

Porcine islet xenografts have the potential to provide an inexhaustible source of islets for ß cell replacement. Proof-of-concept has been established in nonhuman primates. However, significant barriers to xenoislet transplantation remain, including the poorly understood instant blood-mediated inflammatory reaction and a thorough understanding of early xeno-specific immune responses. A paucity of data exist comparing xeno-specific immune responses with alloislet (AI) responses in primates. We recently developed a dual islet transplant model, which enables direct histologic comparison of early engraftment immunobiology. In this study, we investigate early immune responses to neonatal porcine islet (NPI) xenografts compared with rhesus islet allografts at 1 hour, 24 hours, and 7 days. Within the first 24 hours after intraportal infusion, we identified greater apoptosis (caspase 3 activity and TUNEL [terminal deoxynucleotidyl transferase dUTP nick end labeling])-positive cells) of NPIs compared with AIs. Macrophage infiltration was significantly greater at 24 hours compared with 1 hour in both NPI (wild-type) and AIs. At 7 days, IgM and macrophages were highly specific for NPIs (α1,3-galactosyltransferase knockout) compared with AIs. These findings demonstrate an augmented macrophage and antibody response toward xenografts compared with allografts. These data may inform future immune or genetic manipulations required to improve xenoislet engraftment.

Fatal SV40-associated pneumonia and nephropathy following renal allotransplantation in rhesus macaque.

Recrudescence of latent and dormant viruses may lead to overwhelming viremia in immunosuppressed hosts. In immunocompromised hosts, Simian virus 40 (SV40) reactivation is known to cause nephritis and demyelinating central nervous system disease. Here, we report SV40 viremia leading to fatal interstitial pneumonia in an immunosuppressed host following renal allotransplantation.

Selective Targeting of High-Affinity LFA-1 Does Not Augment Costimulation Blockade in a Nonhuman Primate Renal Transplantation Model.

Costimulation blockade (CoB) via belatacept is a lower-morbidity alternative to calcineurin inhibitor (CNI)-based immunosuppression. However, it has higher rates of early acute rejection. These early rejections are mediated in part by memory T cells, which have reduced dependence on the pathway targeted by belatacept and increased adhesion molecule expression. One such molecule is leukocyte function antigen (LFA)-1. LFA-1 exists in two forms: a commonly expressed, low-affinity form and a transient, high-affinity form, expressed only during activation. We have shown that antibodies reactive with LFA-1 regardless of its configuration are effective in eliminating memory T cells but at the cost of impaired protective immunity. Here we test two novel agents, leukotoxin A and AL-579, each of which targets the high-affinity form of LFA-1, to determine whether this more precise targeting prevents belatacept-resistant rejection. Despite evidence of ex vivo and in vivo ligand-specific activity, neither agent when combined with belatacept proved superior to belatacept monotherapy. Leukotoxin A approached a ceiling of toxicity before efficacy, while AL-579 failed to significantly alter the peripheral immune response. These data, and prior studies, suggest that LFA-1 blockade may not be a suitable adjuvant agent for CoB-resistant rejection.

Axial quantitative ultrasound assessment of pediatric bone quality in eastern Nepal.

UNLABELLED: This study presents quantitative ultrasonography (QUS) bone quality data for an underrepresented, south Asian pediatric population from Nepal. Data were collected as part of a longitudinal study of growth and development. This study offers normative data and documents the effect of stunting, wasting, and underweight on the bone properties measured by QUS. INTRODUCTION: The purpose of this study was to (1) examine the bone quality of a rural, non-Western pediatric population using QUS, (2) explore variation in the trajectory of bone quality development between males and females, and (3) examine the impact of growth disruption(s) on bone quality. METHODS: A cross-sectional study of 860 children and adolescents aged 5-18 years from the Jirel ethnic group in eastern Nepal was performed. The Sunlight Omnisense 7000P was used to assess bone quality of the distal 1/3 radius and midshaft tibia. WHO reference standards were used to assess growth disruptions of height, weight, and BMI. RESULTS: QUS bone quality data for an underrepresented, non-Western pediatric population are presented for the radius and tibia. A sizable portion of the study participants were classified as stunted, wasted, and/or underweight. Despite this prevalence of growth disruption in the study sample, bone quality data conform to other documented populations with less growth disruption. Thus, this study offers normative data and documents the minimal effect of stunting, wasting, and underweight on the bone properties measured by QUS. CONCLUSIONS: Non-Western pediatric populations are significantly underserved with regard to simple, non-invasive screening tools that may help identify developmental disorders and assess bone health. The children and adolescents examined here represent normal growth and development for an underrepresented south Asian population. While this work demonstrates that stunting, wasting, or underweight status at time of QUS assessment is not associated with poor bone quality, we do suggest that further study is needed to examine possible cumulative effects of persistent disruptions that may lead to compromised bone quality in later adolescence.

Rescue administration of a helper-dependent adenovirus vector with long-term efficacy in dogs with glycogen storage disease type Ia.

Glycogen storage disease type Ia (GSD-Ia) stems from glucose-6-phosphatase (G6Pase) deficiency and causes hypoglycemia, hepatomegaly, hypercholesterolemia and lactic acidemia. Three dogs with GSD-Ia were initially treated with a helper-dependent adenovirus encoding a human G6Pase transgene (HDAd-cG6Pase serotype 5) on postnatal day 3. Unlike untreated dogs with GSD-Ia, all three dogs initially maintained normal blood glucose levels. After 6-22 months, vector-treated dogs developed hypoglycemia, anorexia and lethargy, suggesting that the HDAd-cG6Pase serotype 5 vector had lost efficacy. Liver biopsies collected at this time revealed significantly elevated hepatic G6Pase activity and reduced glycogen content, when compared with affected dogs treated only by frequent feeding. Subsequently, the HDAd-cG6Pase serotype 2 vector was administered to two dogs, and hypoglycemia was reversed; however, renal dysfunction and recurrent hypoglycemia complicated their management. Administration of a serotype 2 HDAd vector prolonged survival in one GSD-Ia dog to 12 months of age and 36 months of age in the other, but the persistence of long-term complications limited HDAd vectors in the canine model for GSD-Ia.

Strong Dependence of Atmospheric Feedbacks on Mixed-Phase Microphysics and Aerosol-Cloud Interactions in HadGEM3.

We analyze the atmospheric processes that explain the large changes in radiative feedbacks between the two latest climate configurations of the Hadley Centre Global Environmental model. We use a large set of atmosphere-only climate change simulations (amip and amip-p4K) to separate the contributions to the differences in feedback parameter from all the atmospheric model developments between the two latest model configurations. We show that the differences are mostly driven by changes in the shortwave cloud radiative feedback in the midlatitudes, mainly over the Southern Ocean. Two new schemes explain most of the differences: the introduction of a new aerosol scheme and the development of a new mixed-phase cloud scheme. Both schemes reduce the strength of the preexisting shortwave negative cloud feedback in the midlatitudes. The new aerosol scheme dampens a strong aerosol-cloud interaction, and it also suppresses a negative clear-sky shortwave feedback. The mixed-phase scheme increases the amount of cloud liquid water path (LWP) in the present day and reduces the increase in LWP with warming. Both changes contribute to reducing the negative radiative feedback of the increase of LWP in the warmer climate. The mixed-phase scheme also enhances a strong, preexisting, positive cloud fraction feedback. We assess the realism of the changes by comparing present-day simulations against observations and discuss avenues that could help constrain the relevant processes.

Quantifying center of pressure variability in chondrodystrophoid dogs.

The center of pressure (COP) position reflects a combination of proprioceptive, motor and mechanical function. As such, it can be used to quantify and characterize neurologic dysfunction. The aim of this study was to describe and quantify the movement of COP and its variability in healthy chondrodystrophoid dogs while walking to provide a baseline for comparison to dogs with spinal cord injury due to acute intervertebral disc herniations. Fifteen healthy adult chondrodystrophoid dogs were walked on an instrumented treadmill that recorded the location of each dog's COP as it walked. Center of pressure (COP) was referenced from an anatomical marker on the dogs' back. The root mean squared (RMS) values of changes in COP location in the sagittal (y) and horizontal (x) directions were calculated to determine the range of COP variability. Three dogs would not walk on the treadmill. One dog was too small to collect interpretable data. From the remaining 11 dogs, 206 trials were analyzed. Mean RMS for change in COPx per trial was 0.0138 (standard deviation, SD 0.0047) and for COPy was 0.0185 (SD 0.0071). Walking speed but not limb length had a significant effect on COP RMS. Repeat measurements in six dogs had high test retest consistency in the x and fair consistency in the y direction. In conclusion, COP variability can be measured consistently in dogs, and a range of COP variability for normal chondrodystrophoid dogs has been determined to provide a baseline for future studies on dogs with spinal cord injury.

Mechanical and Thermal Sensory Testing in Normal Chondrodystrophoid Dogs and Dogs with Spinal Cord Injury caused by Thoracolumbar Intervertebral Disc Herniations.

BACKGROUND: Intervertebral disc herniation is a common cause of spinal cord injury (SCI) causing paralysis and sensory loss. Little quantitative information is available on the loss and recovery of sensation in dogs with SCI. OBJECTIVES: To determine whether quantitative sensory testing (QST) can be used to establish thermal and mechanical sensory thresholds in chrondrodystrophoid dogs and compare thresholds among normal dogs and dogs with different grades of SCI. ANIMALS: Thirty-three client-owned chondrodystrophoid dogs: 15 normal and 18 SCI dogs. METHODS: Thermal testing was performed by placing a hot (49°C) and cold (5°C) probe on the dorsal metatarsus and mechanical thresholds were tested using calibrated forceps to apply force to the lateral digit. Stimuli were applied until acknowledged, and response rate, latency, and force applied to response were recorded. Test-retest repeatability was determined by calculating intraclass correlation coefficients. Response rates were compared using logistic regression and thresholds were compared using Kaplan-Meier Survival curves. RESULTS: Testing was feasible with moderate repeatability. Thresholds and response rates were significantly different between normal and SCI dogs for all modalities (P < .001). When dogs were grouped by their clinical grade, each grade was significantly different from normal dogs, and cold stimuli differentiated among all grades. CONCLUSION AND CLINICAL IMPORTANCE: Sensory thresholds can be measured reliably in chondrodystrophoid dogs and are altered by SCI. The differences in sensation among neurologic grades indicate that these techniques can be used to further characterize recovery of SCI dogs.

Correlation of early lymphocyte recovery and progression-free survival after autologous stem-cell transplant in patients with Hodgkin's and non-Hodgkin's Lymphoma.

The importance of the association between early lymphocyte recovery and outcome has not been well studied in autologous stem cell transplantation (ASCT). In this retrospective study, we analyzed 90 consecutive patients with non-Hodgkin's and Hodgkin's lymphoma who underwent ASCT. Patients were divided into two groups: group 1 with absolute lymphocyte count (ALC) on day +15 below the median of 667/mm(3), and group 2 with ALC >or=667/mm(3). The median progression-free survival (PFS), but not overall survival (OS), was significantly longer in group 2 when compared to group 1 (16 months vs not reached P=0.02). Group 2 patients also had significantly shorter hospital stay, received higher CD34(+) cell dose, and had shorter time to neutrophil recovery. Multivariate analysis demonstrated day +15 ALC to be an independent prognostic indicator for PFS, but not OS, while CD34(+) cell dose and the number of pretransplant treatments were better predictors for both PFS and OS. We conclude that higher day +15 ALC may independently predict better PFS after ASCT for lymphoma patients; however, whether this merely reflects faster overall recovery caused by higher infused CD34(+) cell dose and less pretransplant therapy needs further investigation.

Cholesterol conservation in skeletal muscle associated with age- and denervation-related atrophy.

The lipid composition of muscles with age- and denervation-atrophy was assayed in whole muscle and isolated sarcolemma of rats aged 10 and 25 months. Although muscle mass decreased at least 17% during aging, muscle cholesterol concentration rose 15-20%. However, phospholipid concentrations were maintained; therefore, the cholesterol-to-phospholipid ratio increased during aging. Plasma cholesterol levels also rose 35%, but this could account for only about 10% of the age-related difference in muscle cholesterol. Likewise, following denervation (7 days) muscle mass decreased by at least 30%, but muscle cholesterol-to-phospholipid ratio increased; the magnitude of the denervation-induced response was unaffected by aging. To localize the source of cholesterol deposition during aging, these assays were repeated on isolated sarcolemma, sarcoplasmic reticulum, and mitochondria membrane fractions. Neither cholesterol nor phospholipid levels changed significantly with age in any of these major membrane systems. Furthermore, sarcolemmal fluidity, which depends on cholesterol content, did not vary significantly with age. At this level of resolution, it thus appears that skeletal muscle membrane phospholipid composition and fluidity do not change appreciably with age. Elevated cholesterol-to-phospholipid ratios detected in whole muscle may be due to cholesterol deposition in non-myofiber locations.

Changes in acetylcholine receptor distribution and binding properties at the neuromuscular junction during aging.

Junctional and extrajunctional acetylcholine receptors were characterized in diaphragm muscle obtained from mature adult and aged rats. Rhodamine-conjugated alpha-bungarotoxin was used to visualize receptor localization. At this level of resolution, there were no major changes in receptor distribution, and nerve terminals were consistently associated with receptors and vice versa. Specific binding characteristics were assayed by measuring 125I-alpha-bungarotoxin binding. Maximal binding to intact junctional and extrajunctional tissue samples was greater in the older rats. The association rate constant in minced tissue decreased in the older animals. Retardation of the initial rate of toxin binding by d-tubocurarine was described by a two-component nonlinear Hofstee plot; values of Ki were about the same for both age groups, but there was a significant shift towards the low-affinity values in the aged rats. Miniature end-plate currents (m.e.p.c.s.) were recorded under voltage-clamp conditions before and after AChE inhibition. When AChE activity was inhibited m.e.p.c. amplitudes and decay time-constants increased in both age groups. The magnitude of these increases was larger in the older animals. Inhibition of AChE did not affect mean channel open time, which was estimated from spectral analyses of ACH-induced membrane noise. Lipid composition was assayed in whole muscle and isolated sarcolemma. Muscle cholesterol concentration rose 15-20 percent, but phospholipid concentrations were maintained. However, neither cholesterol, phospholipid levels, nor membrane fluidity changed significantly with age in isolated sarcolemmal membrane fractions. These data indicate that the numbers of junctional and extrajunctional receptors increase with age. In the junctional region, this is quite likely due to an expanded field of receptors and not an increased density. This is associated with an increased fraction of receptors with lower binding affinity during aging. These changes apparently are not caused by major changes in membrane fluidity or lipid composition.

Poor mobilization of peripheral blood stem cells is a risk factor for worse outcome in lymphoma patients undergoing autologous stem cell transplantation.

The effect of poor blood stem cells mobilization on the outcome of autologous stem cell transplantation (ASCT) has not been well studied. Our aim is to evaluate poor mobilization as a prognostic factor in lymphoma patients undergoing ASCT. We analyzed 90 consecutive patients with Hodgkin's (HD) and non-Hodgkin's lymphoma (NHL) who underwent ASCT. Poor mobilization was defined as the inability to obtain > or = 1 x 10(6) CD34+ cells/kg ideal body weight with two large volume aphereses. Patients were divided into 2 groups: group 1 = poor mobilizers, and group 2 = good mobilizers. The poor mobilizers received lower median transplant CD34+ cell dose (2 x 10(6) vs. 4.5 x 10(6)/kg for good mobilizers, P = 0.001), were more heavily pretreated (P = 0.01), and required higher number of aphereses for PBSC collection (P = 0.0006). The median progression-free survival (PFS) in groups 1 and 2 was 10 and 41 months (P = 0.04), while the median overall survival (OS) was 38 months and not reached (P = 0.02), respectively. Univariate analysis showed that > or = 3 pre-transplant treatments, CD34+ cell dose < or = 2 x 10(6), elevated LDH before transplant, and poor mobilization were significant prognostic factors for poor PFS, while only the first three were significant for worse OS. Multivariate analysis using these same four factors revealed that number of pre-transplant treatments (HR = 6.03, P = 0.001), CD34+ cell dose (HR = 0.1, P = 0.0007) were the only independent predictive factors for worse overall outcome. In conclusion, our data show that poor mobilization could indicate poor outcome in lymphoma patients undergoing ASCT, however, it is more likely to be a reflection of the heavy pre-transplant therapy and lower CD34+ cell dose re-infused in this group of patients.

Incidental paranasal sinus abnormalities on CT of children: clinical correlation.

The paranasal sinuses were prospectively evaluated by CT, clinical history, and physical examination in infants and children having cranial CT for indications unrelated to upper respiratory inflammation (URI). One hundred and one CT scans were studied, and sinus abnormalities were detected in 18% of patients older than 1 year and without signs or symptoms of URI. When signs and/or symptoms of recent URI were present, the incidence of abnormalities was 31%. Maxillary antral were not identifiable or were opacified in 72% of all infants under 1 year old. Because of the high incidence of sinus abnormalities on CT in children with and without evidence of recent URI, abnormalities should not be ascribed to sinusitis without close clinical correlation.

Synovial cysts of the lumbosacral spine: diagnosis by MR imaging.

Intraspinal synovial or ganglion cysts are uncommon lesions associated with degenerative lumbosacral spine disease. CT usually reveals cystic lesions adjacent to a facet joint, and they may show calcification. MR imaging of four surgically confirmed cases of intraspinal synovial cysts revealed subtle signal changes compared with CSF. Short TR/TE images showed the lesions to be slightly hyperintense in three cases and isointense in one case. Long TR/TE sequences revealed a hyperintense appearance in two cases and a hypointense appearance in the others. A peripheral rim of decreased signal on long TR/TE images probably reflects fine calcification or hemorrhage in the margins of the cysts. The multiplanar and contrast characteristics of MR make this technique well suited to the diagnosis of herniated disk, degenerative facet disease, and synovial cyst.

Performance evaluation of a new quality control dose monitor for radiation therapy.

The performance of a new quality control dose monitor for radiation therapy with respect to precision, stability, temperature coefficient, and radiation damage has been extensively evaluated. The results of routine use at several centers are reported.

High dose refractoriness to the neuromuscular toxicity of dithiobiuret in rats.

Increasing the daily dose of dithiobiuret (DTB) given to rats from 0.5 to 1 to 5 mg/kg shortened the latency to onset of treadmill failure and associated flaccid muscle tone from 7 to 5 to 3 days, respectively. Death generally did not follow treadmill failure for at least 2-3 days. After 3 consecutive days of treatment with 5 mg/kg, gastrocnemius muscle contractions elicited by high frequency trains of nerve stimulation were lower in peak tension and more susceptible to tetanic fade (rapid tension decline in tetanus) than were contractions from control rats. Conversely, rats given 12 mg/kg of DTB for 3 days did not exhibit treadmill failure, flaccid skeletal muscle tone, or tetanic contractile abnormalities. Additional treatment with this dose for 2-3 days was required to produce treadmill failure which was not accompanied by flaccid muscle tone. Refractoriness to development of toxicity with DTB was limited in its spectrum; decreased feed and water intake, weight loss, diuresis, dehydration, chromodacryorrhea, and production of mucoid feces occurred even in the absence of flaccid muscle tone. Tissue distribution of DTB-derived [14C] determined 3 hr after injection of [14C]-DTB (12 mg/kg) was largely unaffected by prior treatment with unlabeled DTB (12 mg/kg/day x 2 days). Conventional microelectrode recording studies using end-plates of extensor digitorum longus muscles indicated that abnormalities occurred in quantal release of acetylcholine (ACh) after 5mg/kg/day but not 12 mg/kg/day of DTB. Specifically, reduced quantal content, increased amplitude and prolonged decay of miniature end-plate potentials were observed. The mechanism by which large daily doses overcome or prevent the expected development of flaccid muscle tone and depressed release of ACh typically associated with treatment with DTB does not involve compensatory increases in quantal release of ACh, or altered distribution of the compound.

Dithiobiuret metabolism in the rat.

Our main objective was to describe the metabolism of dithiobiuret (DTB) in the adult, male rat. Based on the thin-layer chromatographic analysis of urine from animals treated ip with 1 mg/kg of [14C] or [35S] labeled DTB, two pathways for metabolism are proposed. One pathway is reversible and involves the oxidation of DTB to thiuret and the reduction of thiuret back to DTB. The other pathway consists of the desulfuration of DTB to monothiobiuret. The liver appears to desulfurate DTB because DTB-derived [35S] was eliminated from the liver more rapidly (T1/2 = 10 hr) than [14C] (T1/2 = 15 hr). The liver was the only tissue where the elimination kinetics of [35S] and [14C] DTB were different. For all extrahepatic tissues examined and plasma, the elimination of DTB-derived [35S] paralleled that of [14C]. The T1/2 for plasma disappearance of both radiolabeled forms of DTB was approximately 10 hr and the cumulative urinary excretion of DTB-derived [35S] and [14C] was parallel and amounted to about 60% of the dose in 24 hr. DTB-derived radioactivity in urine that co-chromatographed with DTB, monothiobiuret, thiuret and sulfate was quantitated along with that of three uncharacterized metabolites. The presence of these unknown metabolites suggests that DTB metabolism is complex. The present study is the first description of the metabolic fate of DTB in the rat and serves as a starting point for determining whether DTB neurotoxicity is caused by the parent compound or a metabolite.

Antagonism of dithiobiuret toxicity in rats.

Daily administration of dithiobiuret (DTB, 1 mg/kg X 6 days, ip) produced delayed onset muscle weakness in rats as indicated by failure in a treadmill test. In nerve-muscle preparations from DTB-intoxicated rats neuromuscular toxicity was manifested as contractile fatigue during tetanic nerve stimulation. As muscle weakness developed, feed intake decreased and the animals lost body weight. Water intake was not altered during this time, but urine output was increased concomitant with the development of muscle weakness and resulted in a state of negative water balance. Daily administration of d-penicillamine (d-PEN) antagonized DTB-induced treadmill failure in a dose-dependent fashion. A daily dose of d-PEN (1 mMol/kg, ip) that completely antagonized treadmill failure also antagonized the contractile fatigue, reduced feed intake, weight loss and negative water balance caused by DTB administration. In rats already intoxicated with DTB, initiating daily d-PEN treatment or discontinuing further DTB administration, caused the animals to recover normal treadmill performance after a latent period of five days. A single dose of d-PEN (1 mMol/kg, iv) was not effective in reversing treadmill failure or contractile fatigue in rats already intoxicated with DTB. Thus, continuous daily administration of d-PEN was necessary for it to be effective. A single dose of d-PEN (1 m Mol/kg, ip) administered one hr after [14C]-DTB (1 mg/kg, ip) did not affect the plasma and tissue concentrations of DTB-derived radioactivity or their corresponding elimination kinetics. Cumulative urinary and fecal excretion of DTB-derived radioactivity were also unaffected by d-PEN administration as were the relative proportions of DTB and two of its metabolites, monothiobiuret and thiuret, in urine. Other agents that produced dose-dependent antagonism of DTB toxicity were diethyldithiocarbamate, disulfiram, cysteamine and 2,2'-dipyridyl. Considering the chemical and biological properties of DTB and its antagonists, a mechanism of antagonism involving an alteration of the thiol-disulfide and/or divalent metal cation status of motor axon terminals is postulated.

Evaluation of the ability of d-penicillamine to protect rats against the neurotoxicity induced by zinc pyridinethione, acrylamide, 2,5-hexanedione and p-bromophenylacetylurea.

Dietary exposure of rats to three different concentrations of zinc pyridinethione (ZPT; 166, 332, 498 ppm) caused delayed onset failure in a treadmill test and, at the higher concentrations (332 and 498 ppm), death. Daily treatment with d-penicillamine (d-PEN) increased the latency period for treadmill failure and lethality. Comparable levels of toxicity were achieved only after d-PEN treated rats had consumed 2-3 times more ZPT than rats not treated with d-PEN. In contrast to ZPT, administration of d-PEN did not affect the onset of treadmill failure associated with acrylamide, p-bromophenylacetylurea or 2,5-hexanedione. Thus, d-PEN provided protection which was selective for ZPT.

Social loafing and social compensation: the effects of expectations of co-worker performance.

Previous research has suggested that people tend to engage in social loafing when working collectively. The present research tested the social compensation hypothesis, which states that people will work harder collectively than individually when they expect their co-workers to perform poorly on a meaningful task. In 3 experiments, participants worked either collectively or coactively on an idea generation task. Expectations of co-worker performance were either inferred from participants' interpersonal trust scores (Experiment 1) or were directly manipulated by a confederate coworker's statement of either his intended effort (Experiment 2) or his ability at the task (Experiment 3). All 3 studies supported the social compensation hypothesis. Additionally, Experiment 3 supported the hypothesis that participants would not socially compensate for a poorly performing co-worker when working on a task that was low in meaningfulness.