Genetic risk factors for the development of allergic disease identified by genome-wide association.

An increasing proportion of the worldwide population is affected by allergic diseases such as allergic rhinitis (AR), atopic dermatitis (AD) and allergic asthma and improved treatment options are needed particularly for severe, refractory disease. Allergic diseases are complex and development involves both environmental and genetic factors. Although the existence of a genetic component for allergy was first described almost 100 years ago, progress in gene identification has been hindered by lack of high throughput technologies to investigate genetic variation in large numbers of subjects. The development of Genome-Wide Association Studies (GWAS), a hypothesis-free method of interrogating large numbers of common variants spanning the entire genome in disease and non-disease subjects has revolutionised our understanding of the genetics of allergic disease. Susceptibility genes for asthma, AR and AD have now been identified with confidence, suggesting there are common and distinct genetic loci associated with these diseases, providing novel insights into potential disease pathways and mechanisms. Genes involved in both adaptive and innate immune mechanisms have been identified, notably including multiple genes involved in epithelial function/secretion, suggesting that the airway epithelium may be particularly important in asthma. Interestingly, concordance/discordance between the genetic factors driving allergic traits such as IgE levels and disease states such as asthma have further supported the accumulating evidence for heterogeneity in these diseases. While GWAS have been useful and continue to identify novel genes for allergic diseases through increased sample sizes and phenotype refinement, future approaches will integrate analyses of rare variants, epigenetic mechanisms and eQTL approaches, leading to greater insight into the genetic basis of these diseases. Gene identification will improve our understanding of disease mechanisms and generate potential therapeutic opportunities.

Risk factors and outcomes analyses at 36 months of a prospective, randomized, multicenter, trial of daclizumab induction in simultaneous kidney-pancreas transplant recipients.

UNLABELLED: The purpose of this study was to analyze risk factors for acute rejection (AR) and long-term outcomes in simultaneous kidney-pancreas transplant (SKPT) patients enrolled in a prospective, multicenter study of daclizumab (DAC) versus no antibody induction. METHODS: A total of 298 SKPT patients were randomized into three groups and categorized based on an intent to treat analysis, and factors associated with AR and survival were identified using logistic regression and Cox proportional hazards models. RESULTS: There were no differences in patient or allograft survival or rejection rates among the three groups at 36 months follow-up. Delayed (kidney) graft function (DGF) was a risk factor for subsequent kidney AR (odds ratio = 2.79, P = .002). The presence of kidney AR was also a risk factor (hazard ratio [HR] = 3.1, P = .003) for kidney graft loss, whereas risk factors for pancreas graft loss (censored for graft loss within 30 days or death with functioning graft) included pancreas AR (HR = 1.97, P = .012), kidney AR (HR = 1.61, P = .042), CMV serostatus donor +/recipient - (HR = 1.62, P = .026), and HLA-B mismatch (HR = 1.58, P = .01). Kidney graft loss (HR = 5.5, P = .02) was the only predictor of mortality. CONCLUSIONS: At 36 months, no significant differences in outcomes were noted in the three study groups. DGF was the major risk factor for kidney AR, kidney AR was the major risk factor for kidney graft loss, and kidney graft loss was the major determinant of mortality. Prevention of kidney DGF and AR in SKPT recipients may play a pivotal role in optimizing long-term outcomes.

A prospective, randomized, multicenter study evaluating the safety and efficacy of two dosing regimens of daclizumab compared to no antibody induction in simultaneous kidney-pancreas transplantation: results at 3 years.

UNLABELLED: This is a report of outcomes at 36 months of a prospective, multicenter study comparing the safety and efficacy of two dosing regimens of daclizumab with no antibody induction in simultaneous kidney-pancreas transplant (SKPT) patients receiving tacrolimus, mycophenolate mofetil, and prednisone. A total of 298 SKPT patients were randomized into one of three groups: daclizumab 1 mg/kg/dose every 14 days for 5 doses (group 1, n = 107); daclizumab 2 mg/kg/dose for 2 doses (group 2, n = 113); and no antibody induction (group 3, n = 78). There were no differences in baseline characteristics among the three groups, and results were analyzed by an intent-to-treat analysis. The incidence of composite events (acute rejection [AR], any allograft lost, or death) at 3 years was 49%, 43%, and 55% in groups 1, 2, and 3, respectively (P = .278). The cumulative incidences of AR were not statistically different among the three groups (P = .178). The mean time to first AR was delayed in groups 2 (288 days) and 1 (245 days) compared to group 3 (145 days, P = .07). There were no differences in patient or allograft survival rates among the three groups, and the rates of serious adverse events, infections, and hospital readmissions were also comparable. Excellent dual graft function in patients with surviving grafts was observed in all three groups at 3 years. CONCLUSIONS: The alternative 2-dose regimen of daclizumab was as safe and effective as the conventional 5-dose regimen compared to no antibody induction in SKPT patients, but no long-term benefits were noted.

A multicenter analysis of the significance of HLA matching on outcomes after kidney-pancreas transplantation.

The purpose of this study was to determine the influence of HLA matching on outcomes in simultaneous kidney-pancreas transplant (SKPT) recipients in a multicenter trial. From March 1999 to May 2001, a total of 297 SKPT recipients were enrolled in a prospective randomized trial of 2 daclizumab dosing strategies versus no antibody induction in combination with tacrolimus, mycophenolate mofetil, and steroids in SKPT recipients. Subanalyses using both univariate and multivariate models were performed at 1 year to identify factors associated with acute rejection, graft loss, or death. Potential risk factors evaluated were treatment group, African American ethnicity, HLA-A mismatches (MM), HLA-B MM, HLA-DR MM, total HLA MM, surgical technique, cytomegalovirus status of donor and recipient, and delayed graft function (DGF). Univariate analyses revealed that treatment group, HLA-A MM, HLA-B MM, total HLA MM >3, and DGF were significantly associated with acute rejection. These variables were then entered into logistic and Cox regression analyses. HLA-A MM and DGF were the only variables that remained significantly associated with acute rejection in the multivariate model. The relative risk for acute rejection in recipients with HLA-A MM was 1.56 (P = .02). In conclusion, despite contemporary immunosuppression, the degree of HLA MM, particularly HLA-A, and DGF are associated with an increased risk for acute rejection in SKPT recipients at 1 year. Less rejection was noted in patients with 0 MM at all 3 HLA loci and in patients with total HLA-MM <3. However, none of these factors affected short-term patient or graft survival rates.

African-American ethnicity is no longer a risk factor for early adverse outcomes in simultaneous kidney-pancreas transplantation with contemporary immunosuppression.

The influence of ethnicity on the outcome of simultaneous kidney-pancreas transplantation (SKPT) is controversial. The aim of this study was to determine the impact of ethnicity on the major endpoints of a prospective, multicenter, randomized trial of two dosing regimens of daclizumab compared to no-antibody induction in SKPT. A total of 297 patients were randomized into three groups: daclizumab 1 mg/kg/dose every 14 days for five doses (group I, n = 107); daclizumab 2 mg/kg/dose every 14 days for two doses (group II, n = 112), and no-antibody induction (group III, n = 78). All patients received tacrolimus, mycophenolate mofetil, and steroids for maintenance immunosuppression. Thirty-seven patients (12.5%) were African-American (AA) and 260 were non-African-American (NAA). Demographics and transplant characteristics were comparable between AA and NAA patients. At 1 year, no differences were seen in patient survival (97% AA, 96% NAA), kidney graft survival (94% AA, 93% NAA), and pancreas graft survival (84% AA, 85% NAA). Rejection rate and incidence of adverse events were similar between AA and NAA subjects. Kidney graft function was comparable between AA and NAA patients at 1 year; however, mean HgbA1C was higher, C-peptide was lower, and oral hypoglycemic use was more common in AA subjects. Thus, in this prospective multicenter study, AA ethnicity was not associated with an increased risk of early adverse outcomes in SKPT. Follow-up will be required to determine whether long-term outcomes remain equivalent, particularly with regard to pancreas graft function.

The impact of delayed graft function of the kidney on the pancreas allograft in simultaneous kidney-pancreas transplantation.

UNLABELLED: It is unclear whether delayed graft function (DGF) of the kidney has any influence on pancreas graft function following simultaneous kidney-pancreas transplantation (SKPT). A subgroup analysis was conducted using data from a multicenter study to determine the impact of DGF of the kidney on pancreas graft function following SKPT. METHODS: Of the 297 SKPT patients, 24 (8%) had DGF of the kidney, defined as the need for dialysis during the first week posttransplant. Clinical parameters including patient and graft survival, incidence of acute rejection, and pancreas and renal function were compared between patients with and without DGF at 1 week, and at 1, 3, 6, and 12 months posttransplant. RESULTS: Demographic and transplant characteristics were similar between the two groups except for longer kidney and pancreas cold ischemia times, more males, and more primary cytomegalovirus (CMV) exposure in the DGF group (P <.05). No differences were seen in patient and graft survival rates, but the incidence of acute renal rejection was higher in patients with DGF (42%) than in those without DGF (15%, P =.001). More patients with DGF (25%) received oral hypoglycemic agents at 1-year posttransplant than in those without DGF (5%, P <.01). At 1 year, the mean serum creatinine was 1.8 mg/dL and 1.4 mg/dL in patients with and without DGF, respectively (P <.01). CONCLUSIONS: Patients with DGF of the kidney had a higher incidence of acute renal rejection and received oral hypoglycemic agents more often during the first year posttransplant compared to those who did not have DGF following SKPT.

Does surgical technique influence outcomes after simultaneous kidney-pancreas transplantation?

Since 1995, many centers have switched from bladder to enteric drainage of the exocrine secretions in simultaneous kidney-pancreas transplantation (SKPT). Enteric exocrine drainage may be performed with either systemic (systemic-enteric [S-E]) or portal (portal-enteric [P-E]) venous delivery of insulin. Controversy exists regarding the optimal surgical technique. From March 1999 to May 2001, a total of 297 SKPT patients were enrolled into a prospective, multicenter, randomized, open-label, comparative trial of two daclizumab dosing strategies versus no-antibody induction in combination with tacrolimus, mycophenolate mofetil, and steroids in SKPT recipients. Surgical techniques were center specific. A total of 171 patients (58%) underwent SKPT with S-E drainage, 96 (32%) with P-E drainage, and 30 (10%) with systemic-bladder (S-B) drainage. The two groups randomized to daclizumab induction were similar with regard to surgical technique (64% S-E, 25% P-E, 11% S-B drainage). Demographic and transplant characteristics and immunosuppression were similar among the three groups, except that more patients with P-E drainage did not receive antibody induction. At 6 months, no differences were seen in patient and graft survival rates, surgical complications including pancreas thrombosis, rates of rejection or infection, readmissions, and kidney and pancreas allograft function among the three different surgical techniques. The 6-month results of this multicenter study suggest no significant differences in outcomes in SKPT recipients according to surgical technique.

Effect of donor-recipient cytomegalovirus serologic status on outcomes in simultaneous kidney-pancreas transplant recipients.

Diabetic patients undergoing simultaneous kidney-pancreas transplantation (SKPT) may be at high risk for developing cytomegalovirus (CMV) infection. To study this issue, we analyzed 297 SKPT patients enrolled into a multicenter trial of two daclizumab dosing strategies versus no antibody induction in combination with tacrolimus, mycophenolate mofetil, and steroids. Complete donor (D) and recipient (R) CMV serology values were available in 294 cases and were distributed as follows: 86 (29%) D+/R-. Eighty-six (29%) D+R+; 45 (16%) D-/R+; 77 (26%) D-/R-; CMV antiviral prophylaxis was center specific, but 98% of patients received either ganciclovir or acyclovir. No differences existed in demographic or transplant characteristics or immunosuppressive regimens among the four groups except that more African-American SKPT recipients were CMV positive at transplant (P <.001). At 6 months, no differences were seen in patient and graft survival rates (GSR) and the incidence of acute rejection (AR) among the groups. However, the CMV D+/R- group had a significantly higher incidence of CMV infection/disease (14%) than the other groups collectively (4%, P <.05). Most cases of CMV infection/disease occurred greater than 3 months posttransplant when prophylaxis was discontinued. In the D-/R- group, the pancreas GSR was higher (94% vs 86% in the remaining three groups) and the incidence of AR was lower (16% vs 25% in the remaining three groups, both P =.09). Primary CMV exposure remains a major risk factor for CMV infection/disease, but does not have an adverse impact on short-term outcomes. Conversely, protective CMV seronegative matching may have a beneficial effect on outcomes.

One-year outcomes in simultaneous kidney-pancreas transplant recipients receiving an alternative dosing regimen of daclizumab.

UNLABELLED: The purpose of this study was to compare the safety and efficacy of two dosing regimen of daclizumab with no-antibody induction in simultaneous kidney-pancreas transplant (SKPT) recipients receiving tacrolimus, mycophenolate mofetil, and steroids. METHODS: A total of 297 SKPT patients were enrolled into this prospective, multicenter, randomized, open-label study. The patients were randomized into three groups: daclizumab 1 mg/kg/dose every 14 days for five doses (group I, n = 107), daclizumab 2 mg/kg/dose every 14 days for two doses (group II, n = 112), and no-antibody induction (group III, n = 78). RESULTS: There were no differences in baseline characteristics among the three groups, except for a higher proportion of African-Americans in group II. The incidence of composite events (acute rejection, graft loss, or death) at 1 year was 36.4%, 32.7%, and 48.7% for groups I, II, and III, respectively (P <.05, group II vs group III). The incidence of acute rejection was highest in group III (34.6%) compared to groups I and II (22.4% and 22.1%, respectively, P <.05). The mean time to acute rejection was delayed in group II (96 days) compared to 23 days in group I and 20 days in group III (P <.05). The adverse-event profiles were comparable among the three groups, except for a higher incidence of infection and readmissions in group III. CONCLUSIONS: Daclizumab was safe and effective in reducing the incidence of acute rejection when compared to no induction. The alternative two-dose regimen of daclizumab was as effective as the conventional five-dose regimen and is logistically more desirable.

Sticky windows: chemical and biological characteristics of the organic film derived from particulate and gas-phase air contaminants found on an urban impervious surface.

A novel environmentally derived mixture that integrates exposure to atmospherically derived gas- and particle-phase compounds in urban areas-namely, the organic film that develops as a thin layer on urban impervious surfaces-was investigated for its ability to induce gene expression via the aryl hydrocarbon receptor (AhR). The organic film on window glass from 21 sites in downtown Toronto (Ontario, Canada) was found to contain a complex mixture of environmental contaminants typical of urban environments, notably PAHs, n-alkanes, PCBs, organochlorine (OC) pesticides, and polar constituents. Using a stably transfected reporter cell line, we found that the crude extract of organic film induces AhR-dependent gene expression in a dose-dependent fashion. Three subfractions of the crude extract induced significant luciferase expression: nonpolar aromatic > polar aromatic > nonpolar aliphatic. Recombination of the fractions did not lead to recovery of the full activity of the crude extract, which may indicate that some of the compounds lost during fractionation were significant contributors to the induction observed with the crude extract. The interactions between a tonic dose of B[ a]P (10(-7) M) and each of the aromatic fractions were determined to be antagonistic following analysis by the method of isoboles. Our results suggest that organic film makes up a diverse array of compounds active at the AhR and that these compounds may not interact in a strictly additive manner.

Ruminant brain ribonucleases: expression and evolution.

Molecular evolutionary analyses of mammalian ribonucleases have shown that gene duplication events giving rise to three paralogous genes occurred in ruminant ancestors. One of these genes encodes a ribonuclease identified in bovine brain. A peculiar feature of this enzyme and orthologous sequences in other ruminants are C-terminal extensions consisting of 17-27 amino acid residues. Evidence was obtained by Western blot analysis for the presence of brain-type ribonucleases in brain tissue not only of ox, but also of sheep, roe deer and chevrotain (Tragulus javanicus), a member of the earliest diverged taxon of the ruminants. The C-terminal extension of brain-type ribonuclease from giraffe deviates much in sequence from orthologues in other ruminants, due to a change of reading frame. However, the gene encodes a functional enzyme, which could be expressed in heterologous systems. The messenger RNA of bovine brain ribonuclease is not only expressed at a high level in brain tissue but also in lactating mammary gland. The enzyme was isolated and identified from this latter tissue, but was not present in bovine milk, although pancreatic ribonucleases A and B could be isolated from both sources. This suggests different ways of secretion of the two enzyme types, possibly related to structural differences. The sequence of the brain-type RNase from chevrotain suggests that the C-terminal extensions of ruminant brain-type ribonucleases originate from deletions in the ancestral DNA (including a region with stop codons), followed by insertion of a 5-8-fold repeated hexanucleotide sequence, coding for a proline-rich polypeptide.

On-line monitoring of wastewater using ion chromatography.

Ion Chromatography (IC) has been used for the on-line determination of anions and cations in a variety of process streams. On-line monitoring of process and wastewater streams optimizes the control of treatment methods by providing early indications of problems that could increase discharges of hazardous compounds to the environment. It is important for the immediate detection and remediation of process upsets in critical streams. The waste flow to the Radioactive Liquid Waste Treatment Facility at the Los Alamos National Laboratory (LANL) is processed before discharge and requires monitoring. Process chromatography is used to monitor the trends of contaminants in real time. The purpose of this study is to develop an automated on-line IC procedure for the simultaneous determination of anions in LANL wastewater.

Determination of inorganic cations in brine solutions by ion chromatography.

The method for analysis of inorganic cations in brine solutions was developed. Ion chromatography is a well-established and accepted technique in the determination of a variety of inorganic ions. However, there are significant complications when ion chromatography is used to determine trace concentrations of inorganic ions in brine matrices. The brine solution in our study was made to simulate the solution from the Waste Isolation Pilot Plant. Instrumental parameters such as eluent composition, flow-rates, and sample loop volumes were investigated to arrive at the optimum condition for the determination of the cations with minimal dilution. Separation was carried out in a Dionex CG12A/CS12A with 8.25 mM H2SO4 as eluent at 1.2 ml/min flow-rate. Our results indicated that ion chromatography is an accurate and a good alternative method for the analysis of cations in brine solution.

Laparoscopic lymphocelectomy: a multi-institutional analysis.

PURPOSE: Because symptomatic lymphoceles are infrequent, single center studies generally report small numbers of patients. We report a multi-institutional experience with and long-term outcome following laparoscopic lymphocelectomy in 81 patients. MATERIALS AND METHODS: Data were obtained from 9 institutions at which at least 5 cases of laparoscopic lymphocelectomy had been performed. Baseline patient demographics, operative time and blood loss, special operative adjunct techniques, postoperative course, convalescence, complications and lymphocele recurrence data were collected and analyzed. RESULTS: A total of 56 men and 25 women with a mean age of 41 years were included in the study. Lymphocele formed after renal transplantation in 78 patients (96%) and after pelvic lymph node dissection in 3 (4%). Average operating time was 123 minutes with a mean blood loss of 43 ml. Omentopexy was performed in 11 cases (13.6%). No intraoperative stenting of the transplant ureter was performed. Intraoperative complications consisted of laryngospasm, bladder injury, inferior epigastric artery injury and mild renal capsule hematoma in 1 patient each. Conversion to open surgery was required for repair of bladder injury in 1, repair of preexisting hernia in 1, unusually thickened lymphocele wall in 1 and inaccessible lymphocele location in 4 cases. Mean time to ambulation and resumption of regular diet was 1 day, and mean hospital stay was 1.5 days. Postoperative complications included trocar site hernia in 1 and urinary retention in 2. Convalescence averaged 2.5 weeks. During a mean followup of 27 months 5 patients (6%) had lymphocele recurrence. CONCLUSIONS: Laparoscopic lymphocelectomy is safe, minimally invasive and effective. It is an excellent alternative to the conventional open surgical approach.

Prednisone withdrawal in kidney transplant recipients on cyclosporine and mycophenolate mofetil--a prospective randomized study. Steroid Withdrawal Study Group.

BACKGROUND: Prospective randomized trials have shown a reduced rate of acute rejection (AR) in mycophenolate mofetil-treated kidney transplant recipients. We hypothesized that this increased protection from AR could allow successful prednisone (P) withdrawal in cyclosporine/mycophenolate mofetil/P-treated recipients. METHODS: A multicenter, prospective, randomized, double-blind trial of P withdrawal at 3 months post-transplant was initiated. Entry criteria were: primary transplant, adult, no AR by 90 days, mycophenolate mofetil dose > or =2 g/day, cyclosporine dose = 5-15 mg/kg/ day, P dose = 10-15 mg/day. Study participants were randomized to have P tapered over 8 weeks (beginning at 3 months posttransplant) to 0 vs. 10 mg/day. Prestudy power analysis determined 500 recipients should be randomized for 80% statistical power to test equivalence of the primary endpoint, AR, or treatment failure at 1 year posttransplant. By design, the study was to be stopped if interim data precluded reaching equivalence. An established data safety monitoring board monitored the study. RESULTS: After 266 patients were enrolled, the patient enrollment was stopped (after safety monitoring board review) because of excess rejection in the P withdrawal group. The Kaplan-Meier estimate of the cumulative incidence of rejection or treatment failure within 1 year posttransplant (+/-95% confidence interval) for the maintenance group was 9.8% (4.4%; treatment failure, 14.9%); for the withdrawal group, 30.8% (21.0%; 39.3%). Treatment differences in the distribution of time to event were highly significant (P = 0.0007). Of note, risk was higher in blacks (39.6%) versus nonblacks (16.0%) (P<0.001). At 1 year post-transplant, there was no difference between groups in patient or graft survival. For the patients with functioning grafts at 6 months posttransplant, withdrawal patients had lower cholesterol (P = 0.0005), had higher creatinine (P = 0.03), and were less likely to use antihypertensives (P = 0.001). These differences persist to 1 yr posttransplant. CONCLUSIONS: We conclude that for recipients on cyclosporine/mycophenolate mofetil/P with no AR at 90 days, the chance of developing subsequent AR is small; if P is tapered and withdrawn, the risk increases (but the majority remain free of acute and chronic rejection). After withdrawal, the risk of AR is different for blacks versus nonblacks. Withdrawal patients had a lower cholesterol level and less need for antihypertensives.

Adenoviral-mediated gene transfer induces sustained pericardial VEGF expression in dogs: effect on myocardial angiogenesis.

OBJECTIVE: Angiogenic peptides like VEGF (vascular endothelial growth factor) and bFGF (basic fibroblast growth factor) have entered clinical trials for coronary artery disease. Attempts are being made to devise clinically relevant means of delivery and to effect site-specific delivery of these peptides to the cardiac tissue, in order to limit systemic side-effects. We characterized the response of the pericardium to delivery of a replication-deficient adenovirus carrying the cDNA for AdCMV.VEGF165, and assessed the effect of pericardial VEGF165 on myocardial collateral development in a canine model of progressive coronary occlusion. METHODS: Ameroid constrictors were placed on the proximal left circumflex coronary artery of mongrel dogs. Ten days later, 6 x 10(9) pfu AdCMV.VEGF165 (n = 9). AdRSV.beta-gal (n = 9), or saline (n = 7) were injected through an indwelling pericardial catheter. Transfection efficiency was assessed by X-gal staining. Pericardial and serum VEGF levels were measured serially by ELISA. Maximal myocardial collateral perfusion was quantified with radiolabeled or fluorescent microspheres 28 days after treatment. RESULTS: In AdRSV.beta-gal-treated dogs, there was extensive beta-gal staining in the pericardium and epicardium, with minimal beta-gal staining in the mid-myocardium and endocardium. Pericardial delivery of AdCMV.VEGF165 resulted in sustained (8-14 day) pericardial transgene expression, with VEGF levels peaking 3 days after infection (> 200 ng/ml) and decreasing thereafter. There was no detectable increase in serum VEGF levels. Maximal collateral perfusion, a principal correlate of collateral development and angiogenesis, was equivalent in all groups. CONCLUSION: Adenoviral-mediated gene transfer is capable of inducing sustained VEGF165 expression in the pericardium; however, locally targeted pericardial VEGF delivery failed to improve myocardial collateral perfusion in this model.