Resolution of low-grade proteinuria is associated with improved outcomes after renal transplantation-a retrospective longitudinal study.

Low-grade proteinuria and systolic hypertension (SHT) are risk factors for allograft failure. Both are dynamic variables and their relationship is not independent. We have simultaneously analyzed the effects of proteinuria and SHT on graft outcomes in 805 adult Kidney Transplant Recipients and impact of their changes over time. Proteinuria and systolic blood pressure (SBP) were recorded for years 1 and 3 posttransplantation. Subjects with proteinuria >1 g/day were excluded. Patients were divided into groups based on proteinuria (Absent(A) <150 mg/day or low-grade(P)150 mg-1 g/day) and blood pressure (Normotensive-SBP <140 mmHg or hypertensive-SBP ≥ 140 mmHg). Graft survival was assessed in all four groups over 10 years by multivariate analysis. At the three annual time points (Year 1, 2 and 3) hypertensive patients with proteinuria had the worst graft survival. Patients with persistent proteinuria between years 1-2 and 2-3 had the poorest graft survival with an improvement if proteinuria regressed (P-A), especially in the Hypertensive group. The impact of proteinuria was highest in persistently hypertensive patients between years 1-3. Thus both proteinuria and SHT were associated with poor graft survival and the combination of the two led to the worst outcomes. Importantly, SHT was associated with significantly worse outcomes in patients with proteinuria. Patient cohort with SHT and low-grade proteinuria represent a selective group that might benefit from intervention.

Enhanced differentiation of osteoclasts from mononuclear precursors in patients with Gaucher disease.

Gaucher disease (GD) is an autosomal recessive disorder caused by deficiency of ß-glucocerebrosidase. Storage of glucosylceramide in reticuloendothelial cells results in multiorgan pathology including bone disease. Established skeletal disease may remain problematic despite Gaucher-specific treatment. Both osteopenia and osteonecrosis have been described but the underlying pathophysiology, in particular the role of monocyte-derived osteoclasts is not well defined. The objective of this study was to explore the effect of glucocerebrosidase deficiency, inhibition and replacement on osteoclast development and function. In cultures derived from GD patients, or where GBA was chemically inhibited multinucleate giant cells expressing markers of osteoclast differentiation occurred earlier and in greater numbers compared to normal controls and the functional capacity of osteoclasts for bone resorption was enhanced. Increases in osteoclast number and activity correlated with radiological markers of active bone disease. Abnormalities were reversed by addition of specific therapies and were attenuated by co-culture with cells derived from healthy controls (HCs). Numbers of osteoblast lineage cells in the peripheral blood were mismatched to osteoclast precursors indicating uncoupling of osteoblast-osteoclast regulation which may further impact on bone remodelling. Elucidation of the underlying mechanisms of these changes will suggest rational therapies for the most disabling aspect of this condition.

An analysis of lymphocyte phenotype after steroid avoidance with either alemtuzumab or basiliximab induction in renal transplantation.

Several studies have analyzed the phenotype of repopulated T-lymphocytes following alemtuzumab induction; however there has been less scrutiny of the reconstituted B-cell compartment. In the context of a randomized controlled trial (RCT) comparing alemtuzumab induction with tacrolimus monotherapy against basiliximab induction with tacrolimus and mycophenolate mofetil (MMF) therapy in renal transplantation, we analyzed the peripheral B- and T-lymphocyte phenotypes of patients at a mean of 25 +/- 2 months after transplantation. We examined the relationship between peripheral lymphocyte phenotype and graft function. Patients who received alemtuzumab had significantly higher numbers of B cells including naïve, transitional and regulatory subsets. In contrast, the CD4(+) T-cell compartment was dominated by a memory cell phenotype. Following either basiliximab or alemtuzumab induction patients with lower numbers of B cells or B subsets had significantly worse graft function. For alemtuzumab there was also a correlation between these subsets the stability of graft function and the presence of HLA-specific antibodies. These results demonstrate that a significant expansion of regulatory type B cells is associated with superior graft function and that this pattern is more common after alemtuzumab induction. This phenomenon requires further prospective study to see whether this phenotype could be used to customize immunotherapy.

Pathology and viral antigen distribution of lethal pneumonia in domestic cats due to pandemic (H1N1) 2009 influenza A virus.

A novel swine-origin H1N1 influenza A virus has been identified as the cause of the 2009 influenza pandemic in humans. Since then, infections with the pandemic (H1N1) 2009 influenza virus have been documented in a number of animal species. The first known cases of lethal respiratory disease associated with pandemic (H1N1) 2009 influenza virus infection in house pets occurred in domestic cats in Oregon. A 10-year-old neutered domestic shorthair and an 8-year-old spayed domestic shorthair died shortly after developing severe respiratory disease. Grossly, lung lobes of both cats were diffusely firm and incompletely collapsed. Histologically, moderate to severe necrotizing to pyonecrotizing bronchointerstitial pneumonia was accompanied by serofibrinous exudation and hyaline membranes in the alveolar spaces. Influenza A virus was isolated from nasal secretions of the male cat and from lung homogenate of the female cat. Both isolates were confirmed as pandemic (H1N1) 2009 influenza virus by real-time reverse transcriptase polymerase chain reaction. With immunohistochemistry, influenza A viral antigen was demonstrated in bronchiolar epithelial cells, pneumocytes, and alveolar macrophages in pneumonic areas. The most likely sources of infection were people in the household with influenza-like illness or confirmed pandemic (H1N1) 2009 influenza. The 2 cases reported here provide, to the best of the authors' knowledge, the first description of the pathology and viral antigen distribution of lethal respiratory disease in domestic cats after natural pandemic (H1N1) 2009 influenza virus infection, probably transmitted from humans.

Patterns of chromosomal evolution in Sigmodon, evidence from whole chromosome paints.

Of the superfamily Muroidea (31 genera, 1578 species), the Sigmodontinae (74 genera, 377 species) is the second largest subfamily in number of species and represents a significant radiation of rodent biodiversity. Only 2 of the 74 genera are found in both North and South America (Sigmodon and Oryzomys) and the remainder are exclusively from South America. In recent molecular studies, the genus Sigmodon (Cricetidae, Sigmodontinae) has been considered sister to many other South American Sigmodontines [Steppan et al., 2004]. We examine the chromosomal evolution of 9 species of Sigmodon utilizing chromosomal paints isolated from S. hispidus, proposed to be similar to the ancestral karyotype [Elder, 1980]. Utilizing a phylogenetic hypothesis of a molecular phylogeny of Sigmodon [Henson and Bradley, 2009], we mapped shared chromosomal rearrangements of taxa on a molecular tree to estimate the evolutionary position of each rearrangement. For several species (S. hirsutus, S. leucotis, S. ochrognathus, S. peruanus, and S. toltecus), the karyotype accumulated few or no changes, but in three species (S. arizonae, S. fulviventer, and S. mascotensis) numerous karyotype rearrangements were observed. These rearrangements involved heterochromatic additions, centric fusions, tandem fusions, pericentric inversions, as well as the addition of interstitial DNA not identified by chromosome paints or C-banding. The hypothesis that the ancestral karyotype for this complex had a diploid number of 52, a fundamental number of 52, and a G-band pattern of which most, if not all are similar to that present in modern day S. hispidus fails to be rejected. This hypothesis remains viable as an explanation of chromosomal evolution in Sigmodontine rodents.

Evidence for biased gene conversion in concerted evolution of ribosomal DNA.

Concerted evolution is the production and maintenance of homogeneity within repeated families of DNA. Two mechanisms--unequal crossing over and biased gene conversion--have been the principal explanations of concerted evolution. Concerted evolution of ribosomal DNA (rDNA) arrays is thought to be largely the result of unequal crossing over. However, concerted evolution of rDNA in parthenogenetic lizards of hybrid origin is strongly biased toward one of two parental sequences, which is consistent with biased gene conversion as the operative mechanism. The apparent gene conversions are independent of initial genome dosage and result in homogenization of rDNA arrays across all nucleolar organizer regions.

Minimal-change nephrotic syndrome due to occupational mercury vapor inhalation.

A 25-year-old man developed nephrotic syndrome and severe hypertension following occupational exposure to mercury vapor whilst working at a fluorescent light factory. A renal biopsy confirmed minimal-change disease on light microscopy, immunofluorescence and electron microscopy. He was also noted to be polycythemic which was initially treated with venesection. His blood and urinary mercury levels were elevated and so he was given chelation therapy with 2,3-dimercaptopropane-1-sulfonate (DMPS), along with steroids for his minimal-change disease, resulting in full resolution of his nephrotic syndrome within 6 weeks. He remains well with normal renal function, blood pressure and normal blood and urine mercury concentrations.

Impact of Single pMOSFET Dielectric Degradation on NAND Circuit Performance.

Degradation of CMOS NAND logic circuits resulting from dielectric degradation of a single pMOSFET using constant voltage stress has been examined by means of a switch matrix technique. As a result, the NAND gate rise time increases by greater than 65%, which may lead to timing errors in high frequency digital circuits. In addition, the NAND gate DC switching point voltage shifts by nearly 11% which may be of consequence for analog or mixed signal applications. Experimental results for the degraded pMOSFET reveal a decrease in drive current by approximately 43%. There is also an increase in threshold voltage by 23%, a decrease in source to drain conductance of 30%, and an increase in channel resistance of about 44%. A linear relationship between the degradation of the pMOSFET channel resistance and the increase in NAND gate rise time is demonstrated, thereby providing experimental evidence of the impact of a single degraded pMOSFET on NAND circuit performance.

Medial-lateral centre of mass displacement and base of support are equally good predictors of metabolic cost in amputee walking.

Amputees are known to walk with greater metabolic cost than able-bodied individuals and establishing predictors of metabolic cost from kinematic measures, such as centre of mass (CoM) motion, during walking are important from a rehabilitative perspective, as they can provide quantifiable measures to target during gait rehabilitation in amputees. While it is known that vertical CoM motion poorly predicts metabolic cost, CoM motion in the medial-lateral (ML) and anterior-posterior directions have not been investigated in the context of gait efficiency in the amputee population. Therefore, the aims of this study were to investigate the relationship between CoM motion in all three directions of motion, base of support and walking speed, and the metabolic cost of walking in both able-bodied individuals and different levels of lower limb amputee. 37 individuals were recruited to form groups of controls, unilateral above- and below-knee, and bilateral above-knee amputees respectively. Full-body optical motion and oxygen consumption data were collected during walking at a self-selected speed. CoM position was taken as the mass-weighted average of all body segments and compared to each individual's net non-dimensional metabolic cost. Base of support and ML CoM displacement were the strongest correlates to metabolic cost and the positive correlations suggest increased ML CoM displacement or Base of support will reduce walking efficiency. Rehabilitation protocols which indirectly reduce these indicators, rather than vertical CoM displacement will likely show improvements in amputee walking efficiency.

Energy flow analysis of amputee walking shows a proximally-directed transfer of energy in intact limbs, compared to a distally-directed transfer in prosthetic limbs at push-off.

Reduced capacity and increased metabolic cost of walking occurs in amputees, despite advances in prosthetic componentry. Joint powers can quantify deficiencies in prosthetic gait, but do not reveal how energy is exchanged between limb segments. This study aimed to quantify these energy exchanges during amputee walking. Optical motion and forceplate data collected during walking at a self-selected speed for cohorts of 10 controls, 10 unilateral trans-tibial, 10 unilateral trans-femoral and 10 bilateral trans-femoral amputees were used to determine the energy exchanges between lower limb segments. At push-off, consistent thigh and shank segment powers were observed between amputee groups (1.12W/kg vs. 1.05W/kg for intact limbs and 0.97W/kg vs. 0.99W/kg for prosthetic limbs), and reduced prosthetic ankle power, particularly in trans-femoral amputees (3.12W/kg vs. 0.87W/kg). Proximally-directed energy exchange was observed in the intact limbs of amputees and controls, while prosthetic limbs displayed distally-directed energy exchanges at the knee and hip. This study used energy flow analysis to show a reversal in the direction in which energy is exchanged between prosthetic limb segments at push-off. This reversal was required to provide sufficient energy to propel the limb segments and is likely a direct result of the lack of push-off power at the prosthetic ankle, particularly in trans-femoral amputees, and leads to their increased metabolic cost of walking.

Significant frequencies of T cells with indirect anti-donor specificity in heart graft recipients with chronic rejection.

BACKGROUND: The purpose of this study was to determine whether T cells with indirect allospecificity could be detected in heart transplant recipients with chronic rejection. METHOD AND RESULTS: Human T-cell clones were used to determine the most effective way to deliver major histocompatibility complex alloantigens for indirect presentation. Seven allograft recipients with evidence of progressive, chronic rejection were selected. Four heart graft recipients with no evidence of chronic rejection were used as controls. Peripheral blood T cells and antigen-presenting cells from the recipients were cultured with frozen/thawed stored donor cells or major histocompatibility complex class I-derived synthetic peptides in limiting dilution cultures and then compared with controls using tetanus toxoid and frozen/thawed third-party cells with no human leukocyte antigens in common with the donor. In 5 of 7 patients analyzed who had chronic rejection, elevated frequencies of T cells with indirect, anti-donor specificity (iHTLf) were detected. No such elevated iHTLf were detected in recipients without chronic rejection. DISCUSSION: iHTLf can be obtained from human transplant recipients, which supports the contention that the indirect pathway is involved in chronic transplant rejection.

Effective sizes and dynamics of uniparentally and diparentally inherited genes.

Models to determine the temporal dynamics and spatial heterogeneity for maternally and paternally inherited genes were derived for populations that may or may not exhibit spatial subdivision. Results were compared to those for diparentally inherited genes. The models permit definition of parameters for mean and variance of litter sizes, breeding group (subpopulation) sizes, and numbers of female mates per male, dispersal rates, and multiple paternity. Exact solutions for asymptotic effective size and spatial divergence (FLS) for maternal and paternal genes are derived. It is shown that solutions for effective size and FLS are transformations of the same quadratic equation. When compared to values for diparentally inherited genes, it is shown that effective sizes for maternal genes may be considerably higher when female dispersal is low as in many mammalian taxa. Likewise, effective sizes for paternal genes may be higher than for diparentally inherited traits when male dispersal is relatively low, as in many species of birds. The traditional assumption that the effective size for maternal genes is approximately equal to the number of females is seldom realized. Spatial heterogeneity and temporal dynamics of genes are inextricably linked as is shown by the interdependency of effective size and spatial heterogeneity.

The end of the LINE?: lack of recent L1 activity in a group of South American rodents.

L1s (LINE-1: Long Interspersed Nuclear Element 1) are present in all mammals examined to date. They occur in both placental mammals and marsupials and thus are thought to have been present in the genome prior to the mammalian radiation. This unusual conservation of a transposable element family for over 100 million years has led to speculation that these elements provide an advantage to the genomes they inhabit. We have recently identified a group of South American rodents, including rice rats (Oryzomys), in which L1s appear to be quiescent or extinct. Several observations support this conclusion. First, genomic Southern blot analysis fails to reveal genus-specific bands in Oryzomys. Second, we were unable to find recently inserted elements. Procedures to enrich for young elements did not yield any with an intact open reading frame for reverse transcriptase; all elements isolated had numerous insertions, deletions, and stop codons. Phylogenetic analysis failed to yield species-specific clusters among the L1 elements isolated, and all Oryzomys sequences had numerous private mutations. Finally, in situ hybridization of L1 to Oryzomys chromosomes failed to reveal the characteristic L1 distribution in Oryzomys with either a homologous or heterologous probe. Thus, Oryzomys is a viable candidate for L1 extinction from a mammalian host.

Isolation of binary species-specific PCR-based markers and their value for diagnostic applications.

Representational difference analysis (RDA), a technique for the isolation of differences between highly similar complex genomes, was employed for isolation of species-specific markers. These markers can be easily adapted for a high throughput PCR-based assay in which multiple specimens can be simultaneously identified based on the presence/absence of amplification products. One of the important features of RDA performed on genomes of different species (interspecific RDA) is its ability to preferentially isolate families of repetitive sequences that are unique to one of the compared genomes. Such families of repetitive DNA are homoplasy-free characters that can be used for cost-efficient, mass identification of specimens in a variety of situations ranging from mark-recapture studies to screenings of egg or larval stages.

Esophageal submucosal gland duct adenoma.

An 81-year-old man with a 3-year history of dysphagia underwent endoscopic resection of a 1-cm-diameter distal esophageal mass. Examination revealed a submucosal neoplasm with a circumscribed growth pattern composed of tubules, cysts, and papillae in association with a marked interstitial lymphoid infiltrate. The cyst lumens and papillae were lined by two to six layers of cytologically bland cuboidal to columnar cells with rare mitotic figures. The basal layer of cells was uniformly positive for smooth-muscle actin. Mucin-positive intracytoplasmic lumens were focally present, but cytoplasmic mucin was not seen. There was no evidence of Barrett's metaplastic epithelium. These features are similar to those in two, possibly three, previously reported cases of esophageal adenomas and bear a resemblance to sialadenoma papilliferum, a rare neoplasm of the minor salivary glands. Their clinicopathologic and immunohistologic features suggest that these neoplasms derive from the submucosal gland ducts. Comparison with the previously reported cases indicates that although the proportions of the various components (tubules, cysts, and papillae) may vary, all cases appear to pursue a slowly growing, clinically indolent course with no evidence of recurrence after complete resection.

N-Arylsulfonylindole derivatives as serotonin 5-HT(6) receptor ligands.

A series of N(1)-arylsulfonyltryptamines were found to be potent ligands of the human serotonin 5-HT(6) receptor with the 5-methoxy-1-benzenesulfonyl analogue (19) having the highest affinity. Additionally, it was discovered that a group such as 3-(3-methoxybenzyl)-1,2,4-oxadiazol-5-yl in the 2-position of the indole ring (43) can replace the arylsulfonyl substituent in the 1-position with no loss of affinity. This suggested that the binding conformation of the aminoethyl side chain at this receptor was toward the 4-position of the indole ring and was supported by the fact that the 4-(aminoethyl)indoles (45) also displayed high affinity, as did the conformationally rigid 1,3,4,5-tetrahydrobenz[c,d]indole (49). Molecular modeling showed that 19, 43, and 45 all had low-energy conformers that overlaid well onto 49. Both 19 and 49 had good selectivity over other serotonin receptors tested, with 49 also showing excellent selectivity over all dopamine receptors. In a functional adenylate cyclase stimulation assay, 19 and 49 had no agonist activity, whereas 45 behaved as a partial agonist. Finally, it was shown that 19 had good activity in the 5-HT(2A) centrally mediated mescaline-induced head twitch assay, which implies that it is brain-penetrant.

The role of the allograft in the induction of donor-specific T cell hyporesponsiveness.

BACKGROUND: With adequate immunosuppression the majority of renal allografts are accepted, despite the exceptional vigour of the T cell alloimmune response. Previous work from this laboratory has demonstrated that this is accompanied by significant reductions in the precursor frequencies of anti-donor T cells. We have also shown that parenchymal cells are tolerogenic in vitro. We propose that the reduction in T cell frequencies may be due to the interaction between circulating T cells and potentially tolerogenic graft parenchymal cells. Primed/memory T cells (CD45RO+) are the only subset capable of reaching the allograft and therefore we would predict that T cell hyporesponsiveness would develop predominantly in the CD45RO+ subset due to their trafficking properties. METHODS: Frequencies of IL-2 secreting CD45RA+ and CD45RO+ CD4+ T cells in response to donor and third party stimulator cells were estimated in a series of renal transplant recipients, both before and after transplantation. RESULTS: There were highly significant reductions in the frequencies of donor-specific CD4+CD45RO+ T cells, when adjusted to control for the generalised effects of immunosuppression. There were no significant alterations in the frequencies of donor-specific CD4+CD45RA+ T cells. CONCLUSIONS: In renal transplant recipients, donor-specific CD4+ T cell hyporesponsiveness occurs predominantly in CD4+ CD45RO+ T cells which is the subset capable of trafficking through the graft.

Biliary scanning with Tc-99m pyridoxylideneglutamate--the effect of food in normal subjects: concise communication.

Technetium-99m pyridoxylideneglutamate biliary scans were performed in 19 normal subjects in both the fasted and nonfasted state. The effect of eating was to reduce visualization of the gallbladder from 100% (fasted) to 47% (nonfasted). The common bile duct was seen in 84% on both occasions but intrahepatic and cystic ducts were seen less frequently in the nonfasted group. Preparation of patients by fasting is essential if information concerning gallbladder function is sought.

Technetium 99m-pyridoxylideneglutamate: a new hepatobiliary radiopharmaceutical. I. Experimental aspects.

The labeling of pyridoxal and the pyridoxylidene derivative of glutamic acid with 99mTc has been achieved by a simple autoclaving procedure. Technetium-99-m-pyridoxylideneglutamate (99mTc-PG) shows marked biliary excretion with accumulation of radioactivity in the gallbladder and intestines of experimental animals. This compound has been extensively investigated with a view to its application in the diagnosis of biliary disorders in man by scintigraphy. Both scintigraphic and quantitative distribution studies showed that 99mTc-PG passed rapidly through the mouse liver with progressive accumulation in the gallbladder, allowing visualization of this organ within 10 min of injection. In 30 min over 40% of the injected dose was excreted into the intestine with an equivalent amount appearing in the urine; however, renal activity remained low. Scintigraphic studies in dogs showed results similar to those obtained in mice. Studies of the toxicity in three animal species indicated a wide margin of safety for 99mTc-PG in the dose proposed for diagnostic purposes in humans.

Technetium-99m calcium phytate--optimization of calcium content for liver and spleen scintigraphy: concise communication.

The addition of ionic calcium to technetium-99m stannous phytate produced an agent with improved splenic uptake. In a series of patients, it was found that molar ratios of 2:1 and 3:1 produced scans of optimal diagnostic quality. Studies in mice showed dose-related changes in the biodistribution. Electron microscopy demonstrated that the addition of calcium produced progressive aggregation of the colloid as the molar ratio was increased. Chemical analysis of sodium phytate was essential to obtain accurate ratios.