Pretransplant HLA typing revealed loss of heterozygosity in the major histocompatibility complex in a patient with acute myeloid leukemia.

INTRODUCTION: Chromosomal abnormalities are frequent events in hematological malignancies. The degree of HLA compatibility between donor and recipient in hematopoietic stem cell transplantation is critical. PURPOSE OF THE STUDY: In this report, we describe an acute myeloid leukemia case with loss of heterozygosity (LOH) encompassing the entire HLA. MATERIALS AND METHODS: HLA molecular typing was performed on peripheral blood (PB) and buccal swabs (BS). Chromosomal microarray analysis (CMA) was performed using a whole genome platform. RESULTS: Typing results on PB sample collected during blast crisis demonstrated homozygosity at the -A, -B, -C, -DR, and -DQ loci. A BS sample demonstrated heterozygosity at all loci. A subsequent PB sample drawn after count recovery confirmed heterozygosity. The CMA performed on PB samples collected during and after blast crisis revealed a large terminal region of copy-neutral LOH involving chromosome region 6p25.3p21.31, spanning approximately 35.9 Mb. The results of the CMA assay on sample collected after count recovery did not demonstrate LOH. CONCLUSIONS: LOH at the HLA gene locus may significantly influence the donor search resulting in mistakenly choosing homozygous donors. We recommend confirming the HLA typing of recipients with hematological malignancies when homozygosity is detected at any locus by using BS samples, or alternatively from PB when remission is achieved.

Prospective Monitoring of Donor-specific Anti-HLA Antibodies After Intestine/Multivisceral Transplantation: Significance of De Novo Antibodies.

BACKGROUND: Presence of circulating donor-specific antibodies (DSA) may be associated with worse clinical outcomes after intestine/multivisceral transplantation. METHODS: In 79 intestine/multivisceral recipients, sera were prospectively screened for DSA by Luminex Single antigen test at 1, 3, 6, 9, 12, 18, 24, and 36 months after transplantation. Standard immunosuppression included thymoglobulin-rituximab induction and tacrolimus-prednisone maintenance. C4d staining was performed retrospectively on biopsies in patients that developed acute rejection (AR). RESULTS: Twenty-two (28%) patients developed de novo DSA at a median posttransplant period of 3 (1-36) months. De novo DSA were observed in 10 of 40 liver-including and 12 of 39 liver-excluding transplants (P = 0.57). Occurrence of AR was slightly higher in patients with de novo DSA (45% vs 33%, respectively; P = 0.41). Similarly, chronic rejection (14% vs 5%; P = 0.21) and graft loss due to AR (18% vs 7%; P = 0.14) were numerically higher in patients with de novo DSA. Only 35% patients experiencing AR had circulating de novo DSA at the time of AR. Antibody-mediated rejection was diagnosed in 6 patients based on C4d staining, of these 2 patients had circulating de novo DSA at the time of biopsy. CONCLUSIONS: De novo DSA formation, particularly early in the posttransplant course may be associated with trends toward worse outcomes. However, its significance in the pathophysiology of AR remains uncertain. Studies focusing mechanisms of DSA-related graft injury and intragraft DSA detection might provide further insight into this issue.

Meningeal relationships to the spinal nerves and rootlets: a gross, histological, and radiological study with application to intradural extramedullary spinal tumors.

INTRODUCTION: Juxtapositional tumors of the spinal nerve roots have been noted to not only interact with the roots at various vertebral levels, but also differ among patients. Therefore, the aim of the current study was to elucidate the potential for variation among the relationships of the meningeal layers at different nerve levels. METHODS: In 20 unembalmed adult cadavers and five fetal specimens, the spinal nerve roots from the cervical, thoracic, and lumbar regions were harvested with their associated meningeal layers and subjected to microdissection, histological analysis, or radiological imaging using 9.4-T MRI. RESULTS: As the nerve rootlets passed from the cord, they received their root sheath covering from the pia. After crossing the subarachnoid space to reach the apertures in the dura, they received two additional looser sheaths, an outer from the dura and an inner from the arachnoid. The pia mater always ended proximal to the arachnoid, and the pia and arachnoid layers extended more distally along the roots with caudal descent. Although the dorsal and ventral roots generally exited through separate dural openings, a single dural opening was also observed, often in the lower spinal regions. Thin intradural septations almost always separated the dorsal and ventral rootlets. The left and right sides frequently differed within individuals. CONCLUSIONS: On the basis of our study, variations of the meninges surrounding the spinal nerve roots are common, but themes do exist. Such data support surgical observations of the different interactions between tumors in these regions with surrounding neural tissues.

Crossmatch-positive liver transplantation in patients receiving thymoglobulin-rituximab induction.

BACKGROUND: Positive crossmatch (CM) in liver transplantation (LT) is associated with worse outcomes. Role of induction immunosuppression in this setting remains to be studied. METHODS: One thousand consecutive LT patients receiving rabbit antithymocyte globulin±rituximab induction were studied. Pretransplantation sera of 55 CM-positive (CM) patients were tested for C1q-fixing donor-specific antibodies (DSA). Diagnosis of antibody-mediated rejection required presence of diffuse vascular C4d expression on liver biopsies. RESULTS: CM was positive in 112 (11%) recipients. Antibody-mediated rejection was observed in 3 (0.03%) patients, whereas acute cellular rejection (ACR) occurred in 31 (3%) patients. CM status was associated with a higher incidence of ACR (9% in CM vs. 2% in CM-negative [CM]; P<0.01) and chronic rejection (4% in CM vs. 1% in CM; P<0.01). Graft survival was slightly lower in CM patients (at 1 year; 85% in CM vs. 89% in CM; P=0.26). Patients with autoimmune hepatitis, primary sclerosing cholangitis, and primary biliary cirrhosis as a group had a tendency toward CM status as well as developing ACR. Upon multivariate analysis, CM status was the strongest predictor of ACR (B=1.14; P=0.02). Only half of CM patients harbored C1q-fixing DSA. Presence of C1q-fixing DSA was not associated with increased incidence of ACR. CONCLUSIONS: In LT, CM status is associated with an increased incidence of acute rejection, chronic rejection, and slightly worse graft survival. With the use of rabbit antithymocyte globulin±rituximab induction, overall low rejection rates can be achieved in CM LT.

The strength of donor-specific antibody is a more reliable predictor of antibody-mediated rejection than flow cytometry crossmatch analysis in desensitized kidney recipients.

The aim of this study was to evaluate the utility of donor-specific antibodies (DSA) and flow cytometry crossmatch (FCCM) as tools for predicting antibody-mediated rejection (AMR) in desensitized kidney recipients. Sera from 44 patients with DSA at the time of transplant were reviewed. Strength of DSA was determined by single antigen Luminex bead assay and expressed as mean fluorescence intensity (MFI). T- and B-cell FCCM results were expressed as mean channel shift (MCS). AMR was diagnosed by C4d deposition on biopsy. Incidence of early AMR was 31%. Significant differences in the number of DSAs (p = 0.0002), cumulative median MFI in DSA class I (p = 0.0004), and total (class I + class II) DSA (p < 0.0001) were found in patients with and without AMR. No significant difference was seen in MCS of T and B FCCM (p = 0.095 and p = 0.307, respectively). The three-yr graft survival in desensitized patients with DSA having total MFI < 9500 was 100% compared to 76% with those having total MFI > 9500 (p = 0.022). Desensitized kidney transplant recipients having higher levels of class I and total DSA MFI are at high risk for AMR and poor graft survival. Recipient DSA MFI appears to be a more reliable predictor of AMR than MCS of FCCM.

Role for a conserved structural motif in assembly of a class I aminoacyl-tRNA synthetase active site.

The catalytic domains of class I aminoacyl-tRNA synthetases are built around a conserved Rossmann nucleotide binding fold, with additional polypeptide domains responsible for tRNA binding or hydrolytic editing of misacylated substrates. Structural comparisons identified a conserved motif bridging the catalytic and anticodon binding domains of class Ia and Ib enzymes. This stem contact fold (SCF) has been proposed to globally orient each enzyme's cognate tRNA by interacting with the inner corner of the L-shaped tRNA. Despite the structural similarity of the SCF among class Ia/Ib enzymes, the sequence conservation is low. We replaced amino acids of the MetRS SCF with portions of the structurally similar glutaminyl-tRNA synthetase (GlnRS) motif or with alanine residues. Chimeric variants retained significant tRNA methionylation activity, indicating that structural integrity of the helix-turn-strand-helix motif contributes more to tRNA aminoacylation than does amino acid identity. In contrast, chimeras were significantly reduced in methionyl adenylate synthesis, suggesting a role for the SCF in formation of a structured active site domain. A highly conserved aspartic acid within the MetRS SCF is proposed to make an electrostatic interaction with an active site lysine; these residues were replaced with alanines or conservative substitutions. Both methionyl adenylate formation and methionine transfer were impaired, and activity was not significantly recovered by making the compensatory double substitution.

Early steroid withdrawal in repeat kidney transplantation.

BACKGROUND AND OBJECTIVES: Kidney re-transplantation (KRT) candidates are considered at high risk for graft failure. Most of these patients are kept on a chronic steroid maintenance (CSM) regimen. The safety of early steroid withdrawal (ESW) remains unanswered in KRT. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: This study was aimed at comparing the outcomes of ESW and CSM in KRT. Retrospective analysis of 113 KRT patients (ESW, n=59; CSM, n=54) was performed. All patients received rabbit anti-thymocyte globulin/steroid induction and were maintained on mycophenolate/tacrolimus (±steroids). RESULTS: One- and 5-year patient survival for the ESW and the CSM group were not significantly different (98 versus 96% and 91 versus 88%, respectively; P=0.991). No significant difference was seen in the graft survival for both groups at 1 and 5 years (98 versus 93% and 80 versus 74%, respectively; P=0.779). Mean 1- and 5-year estimated GFR was not statistically different between the groups (P=0.773 and 0.790, respectively). The incidence of acute rejection at 1 year was 17 and 22% in ESW and CSM patients, respectively (P=0.635). Compared with the ESW group, patients in the CSM group were more likely to be hyperlipidemic (P=0.044), osteoporotic (P=0.010), post-transplant diabetics (P=0.051) and required more medications to control BP (P=0.004). CONCLUSIONS: ESW seems to be a reasonable approach in KRT recipients because the short and intermediate patient survival, graft survival, and graft function is comparable to CSM immunosuppression.

Tolerance induced by anti-CD3 immunotoxin plus 15-deoxyspergualin associates with donor-specific indirect pathway unresponsiveness.

Peritransplant treatment with anti-CD3 immunotoxin plus deoxyspergualin induces tolerance to kidney allografts in most rhesus macaque recipients. Tolerant recipients maintain normal function for years without evidence of chronic rejection. Indirect alloantigen presentation is implicated in chronic rejection. Accordingly, we determined if anti-CD3 immunotoxin plus deoxyspergualin induced rejection-free tolerance associates with suppression of anti-donor indirect pathway responses. Tolerant recipients exhibited an early decrease in direct anti-donor responses with recovery to baseline levels by 3 years posttransplantation. In contrast, tolerant monkeys were unresponsive to donor antigens presented by the indirect pathway. Recipients that rejected their allografts retained vigorous direct and indirect anti-donor responses. Therefore, following temporary donor-specific hyporesponsiveness, direct responses recover in tolerant recipients >1.5 years after transplantation. However, tolerant recipients tested at 1.9-4 years posttransplant are specifically unresponsive to donor antigens presented by the indirect pathway. Thus, the rejection-free state of tolerant recipients may depend on mechanisms regulating indirect pathway responsiveness.