The oral microbiome of patients undergoing treatment for severe aplastic anemia: a pilot study.

The microbiome, an intriguing component of the human body, composed of trillions of microorganisms, has prompted scientific exploration to identify and understand its function and role in health and disease. As associations between microbiome composition, disease, and symptoms accumulate, the future of medicine hinges upon a comprehensive knowledge of these microorganisms for patient care. The oral microbiome may provide valuable and efficient insight for predicting future changes in disease status, infection, or treatment course. The main aim of this pilot study was to characterize the oral microbiome in patients with severe aplastic anemia (SAA) during their therapeutic course. SAA is a hematologic disease characterized by bone marrow failure which if untreated is fatal. Treatment includes either hematopoietic stem cell transplantation (HSCT) or immunosuppressive therapy (IST). In this study, we examined the oral microbiome composition of 24 patients admitted to the National Institutes of Health (NIH) Clinical Center for experimental SAA treatment. Tongue brushings were collected to assess the effects of treatment on the oral microbiome. Twenty patients received standard IST (equine antithymocyte globulin and cyclosporine) plus eltrombopag. Four patients underwent HSCT. Oral specimens were obtained at three time points during treatment and clinical follow-up. Using a novel approach to 16S rRNA gene sequence analysis encompassing seven hypervariable regions, results demonstrated a predictable decrease in microbial diversity over time among the transplant patients. Linear discriminant analysis or LefSe reported a total of 14 statistically significant taxa (p < 0.05) across time points in the HSCT patients. One-way plots of relative abundance for two bacterial species (Haemophilus parainfluenzae and Rothia mucilaginosa) in the HSCT group, show the differences in abundance between time points. Only one bacterial species (Prevotella histicola) was noted in the IST group with a p value of 0.065. The patients receiving immunosuppressive therapy did not exhibit a clear change in diversity over time; however, patient-specific changes were noted. In addition, we compared our findings to tongue dorsum samples from healthy participants in the Human Microbiome Project (HMP) database and found among HSCT patients, approximately 35% of bacterial identifiers (N = 229) were unique to this study population and were not present in tongue dorsum specimens obtained from the HMP. Among IST-treated patients, 45% (N = 351) were unique to these patients and not identified by the HMP. Although antibiotic use may have likely influenced bacterial composition and diversity, some literature suggests a decreased impact of antimicrobials on the oral microbiome as compared to their effect on the gut microbiome. Future studies with larger sample sizes that focus on the oral microbiome and the effects of antibiotics in an immunosuppressed patient population may help establish these potential associations.

Pre-transplant predictors of one yr weight gain after kidney transplantation.

Clinically useful predictors of weight gain could be used to reduce the epidemic of post-kidney transplant obesity and resulting co-morbidities. The purpose of this study was to identify predictors of weight gain at 12 months following kidney transplant in a cohort of 96 recipients. Demographic, clinical, and environmental data were obtained at transplant and 12 months. Descriptive, correlational, and Bayesian network analysis were used to identify predictors. For the 52 (55.9%) recipients who gained weight, the average amount gained was 9.18 ± 6.59 kg. From the 15 baseline factors that met inclusion criteria, Bayesian network modeling identified four baseline predictors for weight gain: younger age, higher carbohydrate consumption, higher trunk fat percentage, and higher perception of mental health quality of life. Three are modifiable through either pre- or immediate post-transplant clinical intervention programs.

Serial analysis of biomarkers of acute pancreas allograft rejection.

Pancreas transplant recipients experience graft loss in spite of improvements in immunosuppressant therapies and diagnostic technologies. Therefore, a method to improve detection and management of acute rejection is needed. This longitudinal study investigated the usefulness of three biomarkers, granzyme B, perforin, and human leukocyte antigen-DR alpha (HLA-DR) measured by real-time PCR on peripheral blood mononuclear cells, for their ability to detect acute rejection and its resolution in 13 recipients of pancreas allograft. Data demonstrated that pre-transplant baseline expression of biomarkers decreased following the initiation of immunosuppression. Throughout follow-up (range 3-27 months), individuals without acute rejection episodes had little variation in their biomarker levels. Recipients with biopsy-proven rejection had a significant increase in the levels of biomarkers as early as five wk before clinical rejection diagnosis. Furthermore, all seven patients with biopsy-proven rejection demonstrated a decrease in the levels of granzyme B and perforin following the increased immunosuppression for the treatment of rejection. This is the first clinical serial measurement of biomarkers in recipients of pancreas transplants. The data demonstrate that upregulation of granzyme B, perforin, and HLA-DR in peripheral blood mononuclear cells are sensitive to changes in the immune environment and could possibly be used to identify those patients at higher risk of rejection.

Blood gene expression profiling in liver transplant recipients with hepatitis C virus and posttransplantation diabetes mellitus.

BACKGROUND: Hepatitis C virus (HCV) is a risk factor for developing posttransplantation diabetes mellitus (PTDM) after liver transplantation; little is known about the biological mechanisms involved with this risk. This study investigated gene expression differences to provide insight into potential mechanisms. PATIENTS AND METHODS: Gene expression profiles of blood samples obtained from 6 HCV+ liver transplant recipients were determined using Affymetrix U133 Plus 2.0 microarrays. Differential gene expression was assessed between HCV+ recipients with PTDM (n = 3) and without PTDM (n = 3) using the GeneSpring 7.3 software package. The Welch t test was used to identify significant differences (P < .05) between groups. Gene expression profiles for 6 HCV- liver transplant recipients (with PTDM = 3, without PTDM = 3) were used as a blind test set to evaluate a subset of genes to predict PTDM. RESULTS: Expression levels of 347 genes were significantly different between recipients with PTDM and those without PTDM. Seventy-four genes were up-regulated and 270 were down-regulated in PTDM. Genes were categorized into functional classes: apoptosis (n = 69 genes); immune function (n = 110); diabetes (n = 17); hepatitis C (n = 12); liver transplant (n = 69). The expression profile of a subset of genes was evaluated for predicting PTDM in 6 HCV- transplant recipients. We accurately predicted the presence or absence of PTDM in 5/6 recipients. CONCLUSIONS: PTDM in HCV+ liver transplant recipients was associated with down-regulated expression of a large number of genes. A subset of these genes was useful to predict PTDM in HCV- recipients. Most genes were associated with apoptosis and immune function. HCV may act as a primer by affecting a group of genes involved in developing diabetes.

Serial peripheral blood cytotoxic lymphocyte gene expression measurements for prediction of pancreas transplant rejection.

Acute rejection after pancreas transplantation remains a significant problem and contributes to immunological graft loss. No clinical markers of pancreas rejection have been universally accepted. The purpose of this study was to investigate the use of genetic markers; granzyme B, perforin, and HLA-DRA in the peripheral blood of pancreas transplant recipients. These genes have been identified in renal and islet cell transplant recipients as noninvasive tools to predict acute rejection. Blood samples were collected weekly for up to 1 year posttransplant. Surveillance biopsies of the pancreas were scheduled at weeks 2, 4, 8, and 12 as part of the typical posttransplant protocol for patients with pancreas alone or pancreas after kidney transplantation. Exclusion criteria included a diagnosis of biopsy-proven chronic rejection alone, pancreatitis, or kidney rejection within 2 months after pancreas biopsy. Gene expression levels of granzyme B, perforin, and HLA-DRA were compared in patients with (n = 7) and without biopsy proven acute rejection (n = 7). Recipients with acute rejection showed increased expression of granzyme B, HLA-DRA, as well as perforin genes compared to patients without biopsy-proven rejection. In addition, we observed that elevation of these genes occurred as early as 4 weeks before the traditional biopsy diagnosis, while the recipients with no rejection showed no change in gene expression. Our data indicated that serial measurements of peripheral blood granzyme B, perforin, and HLA-DRA gene expression can be a useful tool to predict pancreas rejection in its earliest stage.

Gastroparesis following kidney/pancreas transplant.

This pilot study examined associations among patterns of gastric myoelectrical activity, symptoms of gastroparesis, years of diabetes, months of dialysis, and use of gastrointestinal medications in gastroparetic kidney-pancreas (KP) transplant recipients. Electrogastrography (EGG) and gastric symptom data were obtained from 42 transplant recipients before and after transplant (6, 12, and 24 months). Recipients were 38 +/- 7 yr of age, 88% Whites, and 60% male; 97% had hypertension. All had functioning grafts post-transplant (mean creatinine, 1.59 +/- 0.66 mg/dL, and serum glucose 91.97 +/- 24.92 mg/dL). Sixteen subjects had normal EGG (2.7-3.2 cycles per minute, cpm); two were tachygastric (>3.2 cpm) at all time points; one remained bradygastric (<2.7 cpm) throughout the study period. Following transplant, symptoms lessened and were associated with 6-month normalization of EGG (r = 0.41, p = 0.02). A small change in the percentage of patients with normal EGG was observed from baseline to 24 months (67% vs. 69% respectively); however, there was a shift from bradygastria (29% to 15% respectively) to tachygastria (5% to 15% respectively). Prescribed prokinetic and antisecretory medications use increased over the study period from 13 (31%) subjects at baseline to 32 (86%) at 6 months; 21 (78%) at 12 months; and 12 (92%) at 24 months. Although symptoms diminish following transplant, gastroparesis remains a significant problem for transplant patients. Normalization of EGG and shifts from bradygastria to tachygastria occur post-transplant. Our results suggest that serial EGGs and frequent assessment of symptoms can be used to follow gastroparesis in KP recipients.

Race affects insulin and GLP-1 secretion and response to a long-acting somatostatin analogue in obese adults.

OBJECTIVE: This study investigated (1) the effect of octreotide-LAR (Sandostatin-LAR Depot; Novartis) on the enteroinsular axis in a biracial cohort of severely obese adults, (2) whether octreotide suppression of insulin secretion occurs by both a direct beta-cell effect and through mediating a glucagon-like peptide 1 (GLP-1) response, and (3) whether differences in GLP-1 concentrations could explain racial differences in insulin concentrations. DESIGN: Prospective, open-label trial using a pre-post test design. SETTING: Single university, clinical research center. SUBJECTS: In all, 42 healthy, severely obese Caucasian and African-American (AA) adults (93% female, 64% Caucasian, age=37.8+/-1.2 y, weight=123+/-4.2 kg, BMI=44.5+/-1 kg/m(2)), recruited through physician referral and newspaper ads, participated in the study. INTERVENTIONS: Indices of beta-cell activity, insulin and GLP-1 response before and during a 75-gm oral glucose tolerance test were determined before and after 24 weeks of octreotide-LAR. RESULTS: AA exhibited higher beta-cell activity, and insulin and GLP-1 concentrations than Caucasians. Octreotide-LAR suppressed the insulin and GLP-1 levels in both groups.

Racial differences in glucagon-like peptide-1 (GLP-1) concentrations and insulin dynamics during oral glucose tolerance test in obese subjects.

Obese African-American (AA) subjects have higher resting and stimulated insulin concentrations than obese Caucasians (C), which could not be explained by the severity of obesity or the degree of insulin sensitivity. We investigated whether differences in glucagon-like peptide-1 (GLP-1), the most potent incretin that regulates insulin secretion, might explain racial differences in insulin response. Accordingly, we measured fasting and stimulated glucose, insulin, and GLP-1 levels during a 3-h oral glucose tolerance test (OGTT) in 26 obese C (age 38+/-2 y, body mass index 44+/-1 kg/m(2)) and 16 obese AA (age 36+/-2 y, BMI 46+/-2 kg/m(2)) subjects. Corrected insulin response (CIR(30)), a measure of beta-cell activity, whole body insulin sensitivity index (WBISI), and area under the curve (AUC) for insulin, GLP-1, and C-peptide/insulin ratio were computed from the OGTT. Glucose levels, fasting and during the OGTT, were similar between racial groups; 32% of the C and 31% of the AA subjects had impaired glucose tolerance. With a similar WBISI, AAs had significantly higher CIR(30) (2.3+/-0.4 vs 1.01+/-0.1), insulin response (IAUC: 23 974+/-4828 vs 14 478+/-1463), and lower insulin clearance (0.07+/-0.01 vs 0.11+/-0.01) than C (all, P<0.01). Obese AAs also had higher fasting GLP-1 (6.7+/-2.5 vs 4.5+/-1.1) and GLP-1AUC (1174.7+/-412 vs 822.4+/-191) than C (both, P<0.02). Our results indicate that obese AAs had higher concentrations of GLP-1 both at fasting and during the OGTT than obese C. The increased GLP-1 concentration could explain the greater insulin concentration and the increased prevalence of hyperinsulinemia-associated disorders including obesity and type 2 diabetes in AAs.

Posttransplant improvement in heart rate variability correlates with improved quality of life.

A prospective evaluation of 37 kidney and 20 kidney-pancreas transplant recipients was conducted to assess the relationship between pre- to posttransplant changes in heart rate variability (HRV) and quality of life (QoL). Assessments of 24-hour interbeat variability (pNN50 and rMSSD, SDNN, SDANN) and power spectral analysis of total, low (sympathetic), and high (parasympathetic) frequency components of HRV were performed. The Sickness Impact Profile was used to assess three dimensions of QoL (physical, psychosocial, and total functioning) prior to and at 6 months following transplantation. Changes in vagally mediated time domain measures of HRV were related to changes in physical and total functioning. Stronger correlations occurred between biobehavioral measures in kidney-pancreas recipients, with the strongest relationships occurring between changes in HRV frequency domain measures and changes in physical functioning. Findings indicate that changes in HRV and QoL are related, suggesting that interventions that enhance transplant recipients' HRV may also enhance their QoL.

Heart rate variability, mortality, and exercise in patients with end-stage renal disease.

CONTEXT: Cardiac autonomic function has been associated with mortality in patients with end-stage renal disease. It is unknown whether end-stage renal disease patients who have succumbed to sudden cardiac death can be better identified by a newer test of heart rate variability that uses spectral analysis, rather than laboratory evoked measures. OBJECTIVE: This series of studies sought to characterize cardiac autonomic function in patients awaiting kidney transplantation, identify factors associated with heart rate variability, identify tests which distinguish patients at-risk for death, and compare evoked measures with 24-hour heart rate variability measures. PATIENTS: Data were collected on 184 nondiabetics, 60 type 1 diabetics, and 34 type 2 diabetics with end-stage renal disease, all of whom had been referred for kidney transplantation. MAIN OUTCOME MEASURES: The 278 patients and 67 healthy control subjects underwent evoked tests (changes in heart rate with deep breathing and Valsalva maneuver) and 24-hour heart rate variability Holter monitoring (time and frequency domains). Five patients had sudden cardiac deaths during the study. RESULTS: Data showed that end-stage renal disease patients, particularly diabetics, had compromised autonomic function. The standard deviation of all R-to-R intervals for the electrocardiogram recording (< 50 minutes in 60% of the deceased patients), a 24-hour heart rate variability time domain measure, holds the promise of identifying patients at increased risk for death. Exercise was identified as a factor associated with better autonomic function. Examining relationships between 24-hour heart rate variability and characteristics of patients who succumb to death could make quantification of the mortality risk for individual pretransplant end-stage renal disease patients possible, much as it has in other populations. The data from this study may also make it possible to design interventions, such as exercise, aimed at reducing mortality risk.

Changes in patterns of 24-hr heart rate variability after kidney and kidney-pancreas transplant.

BACKGROUND: Transplantation has been shown to improve cardiorespiratory reflex measures of autonomic function. However, there are limited data on how kidney or kidney-pancreas transplantation influence continuous autonomic modulation of heart rate and the clinical utility of 24-hr heart rate variability (HRV) monitoring. METHODS: Ninety nondiabetic kidney and 30 diabetic kidney-pancreas transplant recipients underwent 24-hr Holter monitoring before and again at 6 and 12 months posttransplantation. Tapes were submitted for determination of HRV including interbeat variability (the proportion of adjacent R-R intervals having a difference <50 msec, the SD of all R-R intervals for the entire recording, and the SD of the averages of R-R intervals calculated over 5-min blocks for the entire recording) which is associated with vagal function, sudden death, and circadian function, respectively. Power spectral analysis quantified total neural, sympathetic, and parasympathetic modulation of the heart in ln(msec2). RESULTS: Nondiabetic kidney recipients showed improvement (P< or =0.05) in the SD of the averages of R-R intervals calculated over 5-min blocks (83.2 vs. 95.7 msec) and the SD of all R-R intervals (94.5 vs. 104.4 msec) by 6 months and all groups showed improvement by 12 months. Kidney-pancreas recipients also showed improved total neural (4.35 vs. 4.64) and sympathetic modulation (2.70 vs. 3.13). Kidney-pancreas recipients had significantly poorer values for each measure (P< or =0.05) at all time points. CONCLUSIONS: Cardiac autonomic neuropathy arises in the presence of uremia and diabetes, with severe dysfunction seen when these conditions occur concomitantly. Improvement in cardiac autonomic function follows both kidney and kidney-pancreas transplantation with more pronounced improvement in the circadian measures. Therefore, circadian measures of 24-hr HRV could be used to monitor the restoration of cardiac autonomic function.

Autonomic dysregulation in patients awaiting kidney transplantation.

Sudden cardiac death occurring in patients with end-stage renal disease (ESRD) may be related to poor autonomic function (AF). It is not known whether patients having a sudden death can be identified by commonly used AF evoked tests or if a newer test evaluating heart rate variability (HRV) with power spectral analysis can better distinguish at-risk patients. This study sought to characterize AF in patients awaiting kidney transplantation, to identify factors associated with poor AF and sudden death, and to compare evoked versus 24-hour measures of cardiac AF. All patients underwent evoked cardiac AF tests, which included changes in heart rate with deep breathing (deltaBPM) and valsalva (VR). In addition, 24-hour HRV was assessed with time domain measurements of interbeat variability (pNN50, SDANN, and SDNN), which are associated with vagal function, circadian function, and sudden cardiac death (SDNN < 50), respectively. Frequency domain measures obtained by power spectral analysis (total, low, and high hertz) quantify total neural, sympathetic, and parasympathetic activity of the heart, respectively. Data were collected on 184 nondiabetic patients, 60 type 1 diabetic patients, and 34 type 2 diabetic patients with ESRD referred for transplantation. Five patients, all receiving peritoneal dialysis, experienced nontraumatic sudden cardiac death during the study. Evoked and 24-hour HRV control data were obtained from 67 and 48 healthy adults, respectively. Data show that regardless of subgroup, there was significant AF dysregulation in the 278 patients with ESRD, particularly for those with diabetes and those receiving peritoneal dialysis. Frequency domain measurements (three in each group: nondiabetic patients, type 1 diabetic patients, type 2 diabetic patients, deceased patients, hemodialysis patients, peritoneal dialysis, and nondialysis patients [n = 21]) were most sensitive to dysregulation, with 16 of 21 (76%) measurements more than 2 SD from the mean of the control group. This is in contrast to the time domain measurements (one of 21 [0.04%] > 2 SD from the mean of the control group) and evoked measurements (eight of 14 [57%] outside of the established norms). Of the five deceased patients, only one displayed normal values for all eight AF measurements reported; three (60%) had SDNNs less than 50. Of the 248 surviving patients, 42 (17%) had an SDNN less than 50. When analyzed, the ability of the SDNN to identify an at-risk group was found to have a sensitivity of 60%, a specificity of 83%, a positive predictive value of 7%, a negative predictive value of 99%, and an accuracy of 83%. While AF and time on dialysis were not found to be correlated, the length of diabetes was inversely related to all AF measures (r = -0.27 to -0.48; P < 0.0001), except pNN50. These data suggest that all groups of ESRD patients have severely compromised AF and, regardless of the type of diabetes, those with diabetes have the greatest degree of dysregulation. In addition, individuals receiving peritoneal dialysis were more prone to dysregulation, and the SDNN, a time domain measurement of 24-hour HRV, holds the promise of identifying patients at increased risk for early death in this population.

Cardiovascular dysautonomia of patients with end-stage renal disease and type I or type II diabetes.

BACKGROUND: End-stage renal disease and diabetes mellitus are known to cause autonomic dysfunctions that are responsible for poor outcomes. Studies suggest that 24-hour heart rate variability with power spectral analysis is more sensitive to early changes in autonomic function than laboratory-evoked measures. OBJECTIVES: To evaluate cardiovascular autonomic function in patients (a) awaiting kidney or pancreas-kidney transplantation, (b) without diabetes (NonDM), (c) with Type I insulin dependent diabetes mellitus (IDDM), and (d) with Type II noninsulin dependent diabetes mellitus (NIDDM), and to compare the results of the laboratory-evoked cardiovascular autonomic tests with those from 24-hour heart rate variability monitoring with power spectral analysis. METHOD: This cross-sectional study examined autonomic function in prekidney transplant patients with and without diabetes (N=96), comparing laboratory-evoked measures to 24-hour measures. RESULTS: The nondiabetic group had a normal change in heart rate with deep breathing, Valsalva ratio, and change in systolic blood pressure with tilt. Both diabetic groups had poorer values for all measures of heart rate variability; demonstrated abnormal changes in heart rate with deep breathing and borderline Valsalva ratios; demonstrated a greater decrease in circadian rhythmicity; and had lower SDNNS, pNN50s, and rMSSDs than the nondiabetic group. CONCLUSIONS: Results showed that 24-hour measures are more sensitive, that patients with end-stage renal disease and diabetes regardless of type experience significantly poorer function than do patients without diabetes, and that these values approach those associated with sudden cardiac death.