Anti-Donor HLA Antibody Response After Pancreatic Islet Grafting: Characteristics, Risk Factors, and Impact on Graft Function.

Pancreatic islet grafting restores endogenous insulin production in type 1 diabetic patients, but long-term outcomes remain disappointing as a result of immunological destruction of allogeneic islets. In solid organ transplantation, donor-specific anti-HLA antibodies (DSA) are the first cause of organ failure. This retrospective multicentric study aimed at providing in-depth characterization of DSA response after pancreatic islet grafting, identifying the risk factor for DSA generation and determining the impact of DSA on graft function. Forty-two pancreatic islet graft recipients from the Groupe Rhin-Rhône-Alpes-Genève pour la Greffe d'Ilots de Langerhans consortium were enrolled. Pre- and postgrafting sera were screened for the presence of DSA and their ability to activate complement. Prevalence of DSA was 25% at 3 years postgrafting. The risk of sensitization increased steeply after immunosuppressive drug withdrawal. DSA repertoire diversity correlated with the number of HLA and eplet mismatches. DSA titer was significantly lower from that observed in solid organ transplantation. No detected DSA bound the complement fraction C3d. Finally, in contrast with solid organ transplantation, DSA did not seem to negatively affect pancreatic islet graft survival. This might be due to the low DSA titers, specific features of IgG limiting their ability to activate the complement and/or the lack of allogenic endothelial targets in pancreatic islet grafts.

Identification of a new HLA-A*02 allele, HLA-A*02:305 N, which differs from the HLA-A*02:01:01 allele by a single nucleotide deletion in exon 4.

The HLA-A*02:305N new allele lacks a nucleotide in exon 4 compared to HLA-A*02:01:01 allele.

A new HLA-B*15 allele, B*15:220, found in three individuals sharing the HLA-A*66:01, HLA-C*12:03 and HLADRB1*07:01 alleles.

The new allele HLA-B*15:220 is associated with a conserved haplotype.

Two new HLA-C alleles identified in one volunteer bone marrow donor: C*15:44 and C*07:137:02.

Two new HLA-C alleles were identified in a volunteer bone marrow donor by sequence-based typing.

[Occurrence of severe, persistent thrombocytopenia following allogeneic bone marrow transplantation, attributable to anti-HPA-1 allo-immunisation of the host]. / Survenue d'une thrombopénie sévère et durable, au décours d'une greffe de moelle allogénique, imputable à l'existence chez l'hôte d'une allo-immunisation anti-HPA-1.

A 56 year-old, multiparous woman suffering from a myeloproliferative syndrome, who had received multiple red blood cell and platelet transfusions, was the recipient of an allograft of peripheral blood stem cells derived from her HLA-A, B, DR, DQ and DP and ABO identical sister, following myeloablative conditioning. The persistence of severe, isolated thrombopenia resistant to platelet transfusions led to the discovery of anti-HLA class I allo-immunisation. As HLA compatible platelet transfusions did not result in satisfactory platelet increments, we then discovered the simultaneous presence of anti-HPA-1a allo-immunisation. Genotyping of the HPA-1 systems of the patient (HPA-1B/B) and her sister (HPA-1A/B) enabled us to elucidate the mechanism underlying the persistent thrombopenia and the inefficacy of transfusion. In fact, only transfusion of HPA-1B/B platelets (HLA compatible or incompatible) proved to be efficacious. To reduce the level of anti-HPA-1a antibodies, we performed plasmapheresis sessions and used an anti-CD20 monoclonal antibody. It was only on achieving total haematopoietic chimerism, through rapid interruption of the immunosuppression, that we obtained spontaneous normalisation of the platelet count. The present case emphasises the necessity, before undertaking any allograft of haematopoietic stem cells - even if the latter come from a strictly HLA identical member of the family - of performing a search for eventual anti-HPA allo-immunisation.

Evidence for humoral rejection of a pancreatic islet graft and rescue with rituximab and IV immunoglobulin therapy.

We describe the decline in islet function, in relation to HLA sensitization, in an islet transplant recipient and the recovery of this function after treatment with anti-CD20 monoclonal antibody and IV immunoglobulins. A 51-year-old woman with type 1 diabetes received one intraportal islet infusion. Following this transplantation, she became insulin independent. A search for HLA antibodies by using an ELISA technique remained consistently negative for HLA class I and II. It was only 2 years after the islet transplantation that this search became positive against class II antigens, reaching a peak of reactivity concomitantly with the appearance of a deterioration of glucose control requiring low-dose insulin therapy. Luminex screening and single-antigen assays then revealed the presence of both nondonor-specific and donor-specific antibodies against HLA class II molecules. This immunization, already present in the pretransplant serum, had increased during the 6 months preceding the clinical deterioration. Since these data nevertheless pointed to antibody-mediated rejection of the islet allograft, treatment with anti-CD20 monoclonal antibody and IV immunoglobulins was initiated. One month later, the search by ELISA for antibodies against HLA class II antigens became negative, the Luminex tests normalizing more gradually. As the result of an improvement in glucose control, the patient was again insulin-free.

Second HLA-A*68 null allele, A*6818 N, identified.

A second HLA-A*68 null allele, HLA-A*6818 N, was identified in our laboratory after discrepant results were obtained between class I serological and molecular typing in a male patient suffering from narcolepsy. HLA-A*6818 N displays a sequence identical to that of the HLA-A*6802 allele, except in exon 2 where 20 nucleotides inserted at codon position 48 are a repeat of the 20 preceding nucleotides. This duplication creates a shift of the reading frame, which leads to a premature non-sense codon at position 59 of the null allele.

A null HLA-A*68 allele in a bone marrow donor.

The authors describe an A*68 allele present at the molecular level but not expressed at the cell surface. This non expression results from the deletion of one nucleotide in exon 1, which causes a shift of the reading frame leading to an early non-sense codon in the same exon.

An intronic mutation responsible for a low level of expression of an HLA-A*24 allele.

HLA class I typing performed in parallel by molecular biology and serology has revealed cases where an HLA class I allele was identified but the corresponding antigen on the cell surface was not detected. In the present report, we describe three members of a family in whom an HLA-A24 allele identified at the molecular level was typed as A "blank" by lymphocytotoxicity. This serologically blank antigen was nevertheless faintly detectable by isoelectric focusing (IEF) and FACS analyses. Sequencing of the HLA-A*24 allele from the promoter region to the eighth exonic region revealed a point mutation in the acceptor site of the second intron as compared to the normal HLA-A*24 allele. This mutation could lead to incorrect processing of mRNA through a cryptic acceptor site located at the beginning of the third exon and hence to alternative splicing with a frame shift introducing an early stop codon into the fourth exon.

A nucleotide insertion in exon 4 is responsible for the absence of expression of an HLA-A*01 allele.

HLA class I typing performed in parallel by molecular biology and serology has revealed cases where an HLA class I allele was identified whereas the corresponding antigen was not detected on the cell surface. In the present report, we describe four members of a family in whom an HLA-A1 allele identified at the molecular level was typed as A "blank" by lymphocytotoxicity. This serologically blank antigen was undetectable by isoelectric focusing (IEF). Sequencing of the HLA-A*01 allele from the promoter region to the eighth exonic region revealed insertion of a "C" nucleotide at the beginning of the fourth exon as compared to the common HLA-A*0101 allele. This mutation causes a frame shift, giving rise to an early stop codon in the fourth exon.

A significant percentage of normal T lymphocytes express HLA-DP in the peripheral blood.

HLA-DP expression has been widely investigated on T lymphocytes activated under different conditions. In the present study, a double staining procedure was used in flow cytometric experiments to define DP expression on normal peripheral blood T lymphocytes. In about two-thirds of the case analyzed, DP was expressed on a higher percentage of normal peripheral T lymphocytes than DR was. This was particularly true for 1 of the 16 cases investigated in which the percentage of T lymphocytes expressing DP was 46% and in which DP expression was mainly the prerogative of CD8+ and CD56+ lymphocytes.

A new DR14 allele (DRB1*1411) containing a short DR11 sequence and its haplotypic association.

In the present work, we describe a new DR14 allele. Sequencing of its second DRB1 exon showed it to be DRB1*1404 from codon numbers 9 to 56 and 61 to 86, and DRB1*11 from codons 57 to 60 inclusive.