First-Year Waitlist Hospitalization and Subsequent Waitlist and Transplant Outcome.

Frailty is associated with inferior survival and increased resource requirements among kidney transplant candidates, but assessments are time-intensive and costly and require direct patient interaction. Waitlist hospitalization may be a proxy for patient fitness and could help those at risk of poor outcomes. We examined United States Renal Data System data from 51 111 adult end-stage renal disease patients with continuous Medicare coverage who were waitlisted for transplant from January 2000 to December 2011. Heavily admitted patients had higher subsequent resource requirements, increased waitlist mortality and decreased likelihood of transplant (death after listing: 1-7 days: hazard ratio [HR] 1.24, 95% confidence interval [CI] 1.20-1.28; 8-14 days: HR 1.49, 95% CI 1.42-1.56; ≥15 days: HR 2.07, 95% CI 1.99-2.15; vs. 0 days). Graft and recipient survival was inferior, with higher admissions, although survival benefit was preserved. A model including waitlist admissions alone performed better (C statistic 0.76, 95% CI 0.74-0.80) in predicting postlisting mortality than estimated posttransplant survival (C statistic 0.69, 95% CI 0.67-0.73). Although those with a heavy burden of admissions may still benefit from kidney transplant, less utility is derived from allografts placed in this population. Current kidney allocation policy, which is based in part on longevity matching, could be significantly improved by consideration of hospitalization records of transplant candidates.

Increased Pretransplant Frequency of CD28+ CD4+ TEM Predicts Belatacept-Resistant Rejection in Human Renal Transplant Recipients.

While most human T cells express the CD28 costimulatory molecule constitutively, it is well known that age, inflammation, and viral infection can drive the generation of CD28null T cells. In vitro studies have demonstrated that CD28null cell effector function is not impacted by the presence of the CD28 costimulation blocker belatacept. As such, a prevailing hypothesis suggests that CD28null cells may precipitate costimulation blockade-resistant rejection. However, CD28+ cells possess more proliferative and multifunctional capacity, factors that may increase their ability to successfully mediate rejection. Here, we performed a retrospective immunophenotypic analysis of adult renal transplant recipients who experienced acute rejection on belatacept treatment as compared to those who did not. Intriguingly, our findings suggest that patients possessing higher frequency of CD28+ CD4+ TEM prior to transplant were more likely to experience acute rejection following treatment with a belatacept-based immunosuppressive regimen. Mechanistically, CD28+ CD4+ TEM contained significantly more IL-2 producers. In contrast, CD28null CD4+ TEM isolated from stable belatacept-treated patients exhibited higher expression of the 2B4 coinhibitory molecule as compared to those isolated from patients who rejected. These data raise the possibility that pretransplant frequencies of CD28+ CD4+ TEM could be used as a biomarker to predict risk of rejection following treatment with belatacept.

Belatacept-Resistant Rejection Is Associated With CD28+ Memory CD8 T Cells.

Recently, newer therapies have been designed to more specifically target rejection in an effort to improve efficacy and limit unwanted toxicity. Belatacept, a CD28-CD80/86 specific reagent, is associated with superior patient survival and graft function compared with traditional therapy, but its adoption as a mainstay immunosuppressive therapy has been tempered by increased rejection rates. It is essential that the underlying mechanisms associated with this rejection be elucidated before belatacept is more widely used. To that end, we designed a study in a nonhuman primate kidney transplant model where animals were treated with either a belatacept- or a tacrolimus-based immunosuppressive regimen. Interestingly, we found that elevated pretransplant frequencies of CD28+ CD8+ TEMRA cells are associated with rejection on belatacept but not tacrolimus treatment. Further analysis showed that the CD28+ CD8+ TEMRA cells rapidly lose CD28 expression after transplant in those animals that go on to reject with the allograft infiltrate being predominantly CD28- . These data suggest that CD28+ memory T cells may be resistant to belatacept, capable of further differentiation including loss of CD28 expression while maintaining effector function. The unique signaling requirements of CD28+ memory T cells provide opportunities for the development of targeted therapies, which may synergize with belatacept to prevent costimulation-independent rejection.

Belatacept Combined With Transient Calcineurin Inhibitor Therapy Prevents Rejection and Promotes Improved Long-Term Renal Allograft Function.

Belatacept, a T cell costimulation blocker, demonstrated superior renal function, lower cardiovascular risk, and improved graft and patient survival in renal transplant recipients. Despite the potential benefits, adoption of belatacept has been limited in part due to concerns regarding higher rates and grades of acute rejection in clinical trials. Since July 2011, we have utilized belatacept-based immunosuppression regimens in clinical practice. In this retrospective analysis of 745 patients undergoing renal transplantation at our center, we compared patients treated with belatacept (n = 535) with a historical cohort receiving a tacrolimus-based protocol (n = 205). Patient and graft survival were equivalent for all groups. An increased rate of acute rejection was observed in an initial cohort treated with a protocol similar to the low-intensity regimen from the BENEFIT trial versus the historical tacrolimus group (50.5% vs. 20.5%). The addition of a transient course of tacrolimus reduced rejection rates to acceptable levels (16%). Treatment with belatacept was associated with superior estimated GFR (belatacept 63.8 mL/min vs. tacrolimus 46.2 mL/min at 4 years, p < 0.0001). There were no differences in serious infections including rates of cytomegalovirus or BK viremia. We describe the development of a costimulatory blockade-based strategy that ultimately allows renal transplant recipients to achieve calcineurin inhibitor-free immunosuppression.

The Knife's Edge of Tolerance: Inducing Stable Multilineage Mixed Chimerism but With a Significant Risk of CMV Reactivation and Disease in Rhesus Macaques.

Although stable mixed-hematopoietic chimerism induces robust immune tolerance to solid organ allografts in mice, the translation of this strategy to large animal models and to patients has been challenging. We have previously shown that in MHC-matched nonhuman primates (NHPs), a busulfan plus combined belatacept and anti-CD154-based regimen could induce long-lived myeloid chimerism, but without T cell chimerism. In that setting, donor chimerism was eventually rejected, and tolerance to skin allografts was not achieved. Here, we describe an adaptation of this strategy, with the addition of low-dose total body irradiation to our conditioning regimen. This strategy has successfully induced multilineage hematopoietic chimerism in MHC-matched transplants that was stable for as long as 24 months posttransplant, the entire length of analysis. High-level T cell chimerism was achieved and associated with significant donor-specific prolongation of skin graft acceptance. However, we also observed significant infectious toxicities, prominently including cytomegalovirus (CMV) reactivation and end-organ disease in the setting of functional defects in anti-CMV T cell immunity. These results underscore the significant benefits that multilineage chimerism-induction approaches may represent to transplant patients as well as the inherent risks, and they emphasize the precision with which a clinically successful regimen will need to be formulated and then validated in NHP models.

Fc-Silent Anti-CD154 Domain Antibody Effectively Prevents Nonhuman Primate Renal Allograft Rejection.

The advent of costimulation blockade provides the prospect for targeted therapy with improved graft survival in transplant patients. Perhaps the most effective costimulation blockade in experimental models is the use of reagents to block the CD40/CD154 pathway. Unfortunately, successful clinical translation of anti-CD154 therapy has not been achieved. In an attempt to develop an agent that is as effective as previous CD154 blocking antibodies but lacks the risk of thromboembolism, we evaluated the efficacy and safety of a novel anti-human CD154 domain antibody (dAb, BMS-986004). The anti-CD154 dAb effectively blocked CD40-CD154 interactions but lacked crystallizable fragment (Fc) binding activity and resultant platelet activation. In a nonhuman primate kidney transplant model, anti-CD154 dAb was safe and efficacious, significantly prolonging allograft survival without evidence of thromboembolism (Median survival time 103 days). The combination of anti-CD154 dAb and conventional immunosuppression synergized to effectively control allograft rejection (Median survival time 397 days). Furthermore, anti-CD154 dAb treatment increased the frequency of CD4+ CD25+ Foxp3+ regulatory T cells. This study demonstrates that the use of a novel anti-CD154 dAb that lacks Fc binding activity is safe without evidence of thromboembolism and is equally as potent as previous anti-CD154 agents at prolonging renal allograft survival in a nonhuman primate preclinical model.

Reliability of transpulmonary pressure-time curve profile to identify tidal recruitment/hyperinflation in experimental unilateral pleural effusion.

The stress index (SI) is a parameter that characterizes the shape of the airway pressure-time profile (P/t). It indicates the slope progression of the curve, reflecting both lung and chest wall properties. The presence of pleural effusion alters the mechanical properties of the respiratory system decreasing transpulmonary pressure (Ptp). We investigated whether the SI computed using Ptp tracing would provide reliable insight into tidal recruitment/overdistention during the tidal cycle in the presence of unilateral effusion. Unilateral pleural effusion was simulated in anesthetized, mechanically ventilated pigs. Respiratory system mechanics and thoracic computed tomography (CT) were studied to assess P/t curve shape and changes in global lung aeration. SI derived from airway pressure (Paw) was compared with that calculated by Ptp under the same conditions. These results were themselves compared with quantitative CT analysis as a gold standard for tidal recruitment/hyperinflation. Despite marked changes in tidal recruitment, mean values of SI computed either from Paw or Ptp were remarkably insensitive to variations of PEEP or condition. After the instillation of effusion, SI indicates a preponderant over-distension effect, not detected by CT. After the increment in PEEP level, the extent of CT-determined tidal recruitment suggest a huge recruitment effect of PEEP as reflected by lung compliance. Both SI in this case were unaffected. We showed that the ability of SI to predict tidal recruitment and overdistension was significantly reduced in a model of altered chest wall-lung relationship, even if the parameter was computed from the Ptp curve profile.

Antibody-Mediated Rejection in Sensitized Nonhuman Primates: Modeling Human Biology.

We have established a model of sensitization in nonhuman primates and tested two immunosuppressive regimens. Animals underwent fully mismatched skin transplantation, and donor-specific antibody (DSA) response was monitored by flow cross-match. Sensitized animals subsequently underwent kidney transplantation from their skin donor. Immunosuppression included tacrolimus, mycophenolate, and methylprednisolone. Three animals received basiliximab induction; compared with nonsensitized animals, they showed a shorter mean survival time (4.7 ± 3.1 vs. 187 ± 88 days). Six animals were treated with T cell depletion (anti-CD4/CD8 mAbs), which prolonged survival (mean survival time 21.6 ± 19.0 days). All presensitized animals showed antibody-mediated rejection (AMR). In two of three basiliximab-injected animals, cellular rejection (ACR) was prominent. After T cell depletion, three of six monkeys experienced early acute rejection within 8 days with histological evidence of thrombotic microangiopathy and AMR. The remaining three monkeys survived 27-44 days, with mixed AMR and ACR. Most T cell-depleted animals experienced a rebound of DSA that correlated with deteriorating kidney function. We also found an increase in proliferating memory B cells (CD20(+) CD27(+) IgD(-) Ki67(+) ), lymph node follicular helper T cells (ICOS(+) PD-1(hi) CXCR5(+) CD4(+) ), and germinal center (GC) response. Depletion controlled cell-mediated rejection in sensitized nonhuman primates better than basiliximab, yet grafts were rejected with concomitant DSA rise. This model provides an opportunity to test novel desensitization strategies.

Belatacept: is there BENEFIT for liver transplantation too?

The results of the multicenter belatacept liver transplant trial disappoint with respect to safety and efficacy, and new approaches will be required before this agent plays a role in liver transplant immunosuppression. See article by Klintmalm et al on page 1817.

A novel monoclonal antibody to CD40 prolongs islet allograft survival.

The importance of CD40/CD154 costimulatory pathway blockade in immunosuppression strategies is well-documented. Efforts are currently focused on monoclonal antibodies specific for CD40 because of thromboembolic complications associated with monoclonal antibodies directed towards CD154. Here we present the rational development and characterization of a novel antagonistic monoclonal antibody to CD40. Rhesus macaques were treated with the recombinant anti-CD40 mAb, 2C10, or vehicle before immunization with keyhole limpet hemocyanin (KLH). Treatment with 2C10 successfully inhibited T cell-dependent antibody responses to KLH without significant peripheral B cell depletion. Subsequently, MHC-mismatched macaques underwent intraportal allogeneic islet transplantation and received basiliximab and sirolimus with or without 2C10. Islet graft survival was significantly prolonged in recipients receiving 2C10 (graft survival time 304, 296, 265, 163 days) compared to recipients receiving basiliximab and sirolimus alone (graft survival time 8, 8, 10 days). The survival advantage conferred by treatment with 2C10 provides further evidence for the importance of blockade of the CD40/CD154 pathway in preventing alloimmune responses. 2C10 is a particularly attractive candidate for translation given its favorable clinical profile.

Experience with a novel efalizumab-based immunosuppressive regimen to facilitate single donor islet cell transplantation.

Islet transplantation is an experimental therapy for selected patients with type 1 diabetes (T1DM). It remains limited by immunosuppressive drug toxicity, progressive loss of insulin independence, allosensitization and the need for multiple islet donors. We describe our experience with an efalizumab-based immunosuppressive regimen as compared to the prevailing standard regimen, the Edmonton protocol. Twelve patients with T1DM received islet transplants: eight were treated with the Edmonton protocol; four were treated with daclizumab induction, a 6-month course of tacrolimus, and maintenance with efalizumab and mycophenolate mofetil. The primary endpoint was insulin independence after one islet infusion. Only two Edmonton protocol treated patients achieved the primary endpoint; six required islets from multiple donors, and all experienced leukopenia, mouth ulcers, anemia, diarrhea and hypertransaminasemia. Four became allosensitized. All patients treated with the efalizumab-based regimen achieved insulin independence with normal hemoglobin A1c after a single islet cell infusion and remained insulin independent while on efalizumab. These patients experienced significantly fewer side effects and none became allosensitized. Trial continuation was terminated by withdrawal of efalizumab from the market. These data suggest that this efalizumab-based regimen prevents islet rejection, is well tolerated, and allows for single donor islet transplantation.

Nitrogen leaching from Douglas-fir forests after urea fertilization.

Leaching of nitrogen (N) after forest fertilization has the potential to pollute ground and surface water. The purpose of this study was to quantify N leaching through the primary rooting zone of N-limited Douglas-fir [Pseudotsuga menziesii (Mirb.) Franco] forests the year after fertilization (224 kg N ha(-1) as urea) and to calculate changes in the N pools of the overstory trees, understory vegetation, and soil. At six sites on production forests in the Hood Canal watershed, Washington, tension lysimeters and estimates of the soil water flux were used to quantify the mobilization and leaching of NO(3)-N, NH(4)-N, and dissolved organic nitrogen below the observed rooting depth. Soil and vegetation samples were collected before fertilization and 1 and 6 mo after fertilization. In the year after fertilization, the total leaching beyond the primary rooting zone in excess of control plots was 4.2 kg N ha(-1) (p = 0.03), which was equal to 2% of the total N applied. The peak NO(3)-N concentration that leached beyond the rooting zone of fertilized plots was 0.2 mg NO(3)-N L(-1). Six months after fertilization, 26% of the applied N was accounted for in the overstory, and 27% was accounted for in the O+A horizon of the soil. The results of this study indicate that forest fertilization can lead to small N leaching fluxes out of the primary rooting zone during the first year after urea application.

Mathematical models for pressure controlled ventilation of oleic acid-injured pigs.

One-compartment, mathematical models for pressure controlled ventilation, incorporating volume dependent compliances, linear and nonlinear resistances, are constructed and compared with data obtained from healthy and (oleic acid) lung-injured pigs. Experimental data are used to find parameters in the mathematical models and were collected in two forms. Firstly, the P(e)-V curves for healthy and lung injured pigs were constructed; these data are used to compute compliance functions for each animal. Secondly, dynamic data from pressure controlled ventilation for a variety of applied pressures are used to estimate resistance parameters in the models. The models were then compared against the collected dynamic data. The best mathematical models are ones with compliance functions of the form C(V) = a + bV where a and b are constants obtained from the P(e)-V curves and the resistive pressures during inspiration change from a linear relation P(r) = RQ to a nonlinear relation P(r) = RQ(epsilon) where Q is the flow into the one-compartment lung and epsilon is a positive number. The form of the resistance terms in the mathematical models indicate the possible presence of gas-liquid foams in the experimental data.

Surveys of long-term ventilatory support in Minnesota: 1986 and 1992.

To determine the prevalence and some characteristics of persons in Minnesota receiving long-term ventilatory support (ventilator-assisted individuals [VAIs]), we conducted a survey of this population in Minnesota in 1986 and then again in 1992 by canvassing long-term care units and home medical equipment providers. The number of VAIs in 1992 was 110 percent greater than in 1986 (216 vs 103). In 1986, 81 percent of these patients received care at home; the remaining patients were supported in long-term care facilities. By 1992, the percentage had changed to 65 percent supported in the home and 35 percent in long-term care facilities. In both surveys, the largest number of VAIs were in the diagnostic categories of poliomyelitis, cervical trauma, amyotrophic lateral sclerosis (ALS), chronic obstructive pulmonary disease (COPD), and muscular dystrophy. The primary diagnoses with the greatest increase in number of patients were cervical trauma and ALS. When VAIs were categorized by age groups, there was a large increase in the proportion of patients younger than 10 years of age and older than 60 years of age. While the number of patients is small, the total resources required for care of these patients can be substantial. These data suggest that we need to monitor the number and demographic characteristics of VAIs in the United States so that appropriate policies and programs are developed to provide effective support services.

Estimated and determined P50 values in erythrocytosis.

Actual determination of the position of the oxygen-hemoglobin dissociation curve (P50) was compared to three methods of estimating P50 in 20 patients with erythrocytosis. There was a significant correlation between determined and estimated P50 values for each of the methods studied. Furthermore, four patients with high affinity hemoglobins were readily distinguished from the other patients by estimating P50. The data suggest that calculating P50 is reliable and may replace the need for actual measurement.

Effect of a nasogastric tube on esophageal pressure measurement in normal adults.

We studied the correspondence between fluctuations of esophageal pressure measured before and after placement of a nasogastric (NG) tube in six normal volunteers. Flow, airway pressure, and esophageal pressure data from at least 20 breaths were recorded in seven ventilatory conditions in two body postures: 0 degree (supine) and 60 degrees (upright). The conditions studied included normal quiet breathing, added resistance, reduced compliance, increased frequency, increased tidal volume, continuous positive airway pressure, and volume-cycled ventilation with positive pressure. During recording with the NG tube in place, the subject targeted the same tidal volume (VT), respiratory rate, and inspiratory time fraction (TI/TTOT) recorded before NG tube placement. A computer program selected for analysis only those recorded breaths with and without an NG tube that were "matched" within 5 percent for both VT and TI. We calculated average VT, TI, and esophageal pressure fluctuation (delta Pes) for the matched breaths from each subject during every condition. The delta Pes values with and without NG tube were not statistically different in any tested condition (p > 0.05). Our data indicate that the presence of an NG tube does not invalidate the accuracy of delta Pes measurements made using a well-positioned balloon catheter in the tested conditions.

Spirometry and maximal respiratory pressure references from healthy Minnesota 65- to 85-year-old women and men.

OBJECTIVE: To obtain spirometry and maximal respiratory pressure (MRP) reference values for elderly persons. DESIGN: Survey. SETTING: General community. PARTICIPANTS: Four hundred seventy-one healthy ambulatory white women and men age 65+ years. METHODS: A stringent spirometry quality assurance program exceeded American Thoracic Society recommendations. A "healthy" subgroup of 176 women and 112 men between the ages of 65- and 85 years were identified by excluding those with conditions that negatively influenced FEV1 in a multiple regression analysis. Reference equations and normal ranges for FEV1, FVC, FEF25-75%, peak flow, and maximal inspiratory and expiratory pressures (MRPs) were determined from the healthy group with good quality maneuvers. RESULTS: Less than 10% of the subjects were unable to perform three acceptable spirometry maneuvers and ten MRP maneuvers. When the age and height corrected FEV1s from this group were compared with other spirometry reference studies, mean values from the women were nearly identical to those from Morris, while these men had substantially lower FEV1 values (by 0.3- to 0.5L) than elderly men in Crapo's study. Mean peak flow was over 20% higher when compared with previous studies, suggesting greater initial expiratory effort by our subjects. The maximal inspiratory pressure (MIP) values were about 20% higher than those reported by the Cardiovascular Health Study, perhaps because five MIP maneuvers were always performed. CONCLUSION: Spirometry and MRP reference values used for elderly patients should come from population studies using similar techniques and with large numbers of subjects over age 65 years.

Distal effects of tracheal gas insufflation: changes with catheter position and oleic acid lung injury.

We separated distal (turbulence-related) and proximal (dead space washout-related) effects of tracheal gas insufflation (TGI) by comparing the effects of straight and inverted catheters. We reasoned that the inverted catheter was unlikely to remove CO2 from conducting airways distal to its orifice. In six normal dogs during TGI at 10 l/min, advancing the catheters from 10 to 1 cm above the main carina decreased dead space volume by 29 +/- 12 and 12 +/- 6 ml (P < 0.04) with the straight and inverted catheters, respectively. By comparison, the tracheal volume between 10 and 1 cm above the carina was 15 +/- 2 ml. In another set of dogs (n = 5), we examined the distal effects of TGI before and after oleic acid-induced lung injury. During TGI at 10 l/min before and after oleic acid injury, the differences in arterial PCO2 between the straight and inverted catheters were 5 +/- and 9 +/- 6 Torr (P < 0.18), respectively. Our data suggest that distal effects of TGI become more pronounced as the catheter tip is positioned closer to the main carina. The distal effects of TGI were not diminished after oleic acid injury when minute ventilation was maintained constant.

Comparison of mathematical and mechanical models of pressure-controlled ventilation.

Recent evidence that volume-cycled mechanical ventilation may itself produce lung injury has focused clinical attention on the pressure waveform applied to the respiratory system. There has been an increasing use of pressure-controlled ventilation (PCV), because it limits peak cycling pressure and provides a decelerating flow profile that may improve gas exchange. In this mode, however, the relationships are of machine adjustments to ventilation and alveolar pressure are not straightforward. Consequently, setting selection remains largely an empirical process. In previous work, we developed a biexponential model of PCV that provides a conceptual framework for understanding these interactions (J. Appl. Physiol. 67: 1081-1092, 1989). We tested the validity of this mathematical model in a single-compartment analogue of the respiratory system across wide ranges of clinician-set variables (frequency, duty cycle, applied pressure) and impedance conditions (inspiratory and expiratory resistance and system compliance). Our data confirm the quantitative validity of the proposed model when approximately rectilinear waves of pressure are applied and appropriate values for impedance are utilized. Despite a fixed-circuit configuration, however, resistance proved to be a function of each clinician-set variable, requiring remeasurement of system impedance as adjustments in these variables were made. With further modification, this model may provide a practical as well as a conceptual basis for understanding minute ventilation and alveolar pressure fluctuations during PCV in the clinical setting.