Monitoring indirect allorecognition in renal transplant recipients;(INDIRECT)

Monitoring indirect allorecognition in renal transplant recipients;(INDIRECT) - Indirect allorecognition in renal transplantation

Status

Active, not recruiting

Phases

Unknown

Study type

Observational

Source

NL-OMON

Registry ID

NL-OMON37042

Enrollment

Registered

2012-08-23

Start date

2012-11-29

Completion date

Unknown

Last updated

2024-04-29

For informational purposes only — not medical advice. Sourced from public registries and may not reflect the latest updates. Terms

Conditions

renal transplantation

Interventions

None listed

Eligibility

Age

18 Years to 99 Years

Inclusion criteria

Inclusion criteria: Inclusion criteria for renal transplant patients: • Age: 18 - 80 years • Female or male • First kidney transplantation (living and cadaveric) • Patients must be able to give informed consent and the consent must be obtained prior to any study procedure. • Patients must be transplanted with an HLA-A1, A2, B7 and/or B8 mismatched kidney. (Recipients must therefore have at least one mismatch with one of the previously mentioned HLA-I) • At least 6 months after transplantation

Exclusion criteria

Exclusion criteria: Exclusion criteria for renal transplant patients: • Double organ transplantation. • Patients with a previous transplantation since the earlier mismatch might complicate interpretation. • Patients who had a rejection episode less than 2 months before. • Patients with evidence of active infection or abscesses • Patients suffering from an active autoimmune disease • Subjects who currently have an active opportunistic infection (e.g., herpes zoster [shingles], cytomegalovirus (CMV), Pneumocystis carinii (PCP), aspergillosis, histoplasmosis, or mycobacteria other than TB) • Malignancy (including lymphoproliferative disease) within the past 2-5 years (except for squamous or basal cell carcinoma of the skin that has been treated with no evidence of recurrence).

Design outcomes

Secondary

Measure	Time frame
5.1.2 Secondary study parameters/endpoints (if applicable) Investigate the correlation between frequencies of T cells with indirect reactivity T-cells and simultaneous presence alloreactive antibodies.	—

Primary

Measure	Time frame
5. METHODS 5.1 Study parameters/endpoints 5.1.1 Main study parameter/endpoint Test whether recombinant HLA molecules (A1, A2, B7, B8) can be used for the monitoring of the indirect allorecognition pathway in renal transplant recipients (as described in study procedures) 5.1.3 Other study parameters (if applicable) Not applicable 5.2 Randomisation, blinding and treatment allocation Not applicable 5.3 Study procedures For all patients 50ml (5x 10ml) of heparinized blood will be obtained during one standard visit for blood and urine samples, for the measurement of indirectly reactive T-cells. We will determine in the same blood samples presence of anti-HLA antibodies. Mononuclear cells will be isolated from the 40ml of obtained blood using Ficoll-Hypaque (Pharmacie LUMC, Leiden, the Netherlands). Based on normal values for PBMCs at least 3610^6 cells will be obtained. 110^6 PBMCs/well will be plated in 96-well plate with/without 25µg/ml HLA-I monomer (the mismatched monomer will be added). As a control, cultures will be performed with/without 2µg/ml MHC-II antibodies to inhibit the indirect T-cell response. Remaining activity represent the semi-direct allorecognition and should be considered as the background reactivity level. The PBMCs will be incubated with the HLA-I monomer for 4 hours, after which they will be washed. HLA-II antibodies will then be added for at least 24h at which time supernatants will be collected and cytokines will be measured. Moreover, cultures will be supplemented with 3H thymidine, and after overnight culture, proliferation will be measured. A total volume of 200µl will be used throughout the assay. All tests will be performed at least in triplo and if required culture conditions will be further optimized.	—

Measure

Time frame

5. METHODS 5.1 Study parameters/endpoints 5.1.1 Main study parameter/endpoint Test whether recombinant HLA molecules (A1, A2, B7, B8) can be used for the monitoring of the indirect allorecognition pathway in renal transplant recipients (as described in study procedures) 5.1.3 Other study parameters (if applicable) Not applicable 5.2 Randomisation, blinding and treatment allocation Not applicable 5.3 Study procedures For all patients 50ml (5x 10ml) of heparinized blood will be obtained during one standard visit for blood and urine samples, for the measurement of indirectly reactive T-cells. We will determine in the same blood samples presence of anti-HLA antibodies. Mononuclear cells will be isolated from the 40ml of obtained blood using Ficoll-Hypaque (Pharmacie LUMC, Leiden, the Netherlands). Based on normal values for PBMCs at least 36*10^6 cells will be obtained. 1*10^6 PBMCs/well will be plated in 96-well plate with/without 25µg/ml HLA-I monomer (the mismatched monomer will be added). As a control, cultures will be performed with/without 2µg/ml MHC-II antibodies to inhibit the indirect T-cell response. Remaining activity represent the semi-direct allorecognition and should be considered as the background reactivity level. The PBMCs will be incubated with the HLA-I monomer for 4 hours, after which they will be washed. HLA-II antibodies will then be added for at least 24h at which time supernatants will be collected and cytokines will be measured. Moreover, cultures will be supplemented with 3H thymidine, and after overnight culture, proliferation will be measured. A total volume of 200µl will be used throughout the assay. All tests will be performed at least in triplo and if required culture conditions will be further optimized.

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Countries

Netherlands

Outcome results