Essential genetic modifiers and their measurable impact in a community-recruited population analysis for non-severe hemoglobin E/ß-thalassemia prenatal genetic counseling.

The study aimed to identify essential phenotype-modulating factors among the pre-existence of several important ones and clarify their measurable impact on the clinical severity of hemoglobin (Hb) E/ß-thalassemia in a community-recruited population analysis. This prospective study was designed to compare modifiers between community- (less or no symptoms) and hospital-recruited individuals with Hb E/ß-thalassemia. The formerly included couples previously assessed for prenatal thalassemia at-risk status at 42 community and 7 referral hospitals in Thailand through on-site investigations between June 2020 and December 2021. The control included Hb E/ß-thalassemia patients undergoing transfusions. The Mahidol score classified disease severity. Beta-globin, α0-thalassemia (-SEA, -THAI), α+-thalassemia (-α3.7, -α4.2), Hb Constant Spring (αCS) alleles, rs766432 in BCL11A, rs9399137 in HBS1L-MYB, and rs7482144-XmnI were evaluated. Modifiers were compared between 102 community- and 104 hospital-recruited cases. Alleles of ß+, -SEA, -α3.7, αCS, and a minor allele of rs9399137 were prevalent in the community and mild severity groups (p < 0.05). Multiple linear regression analysis associated modulating alleles with -4.299 (-SEA), -3.654 (ß+), -3.065 (rs9399137, C/C), -2.888 (αCS), -2.623 (-α3.7), -2.361 (rs7482144, A/A), -1.258 (rs9399137, C/T), and -1.174 (rs7482144, A/G) severity score reductions (p < 0.05). Certain modifiers must be considered in routine prenatal genetic counseling for Hb E/ß-thalassemia.

Lessons learned from a prenatal diagnosis program for thalassemia in Thailand.

OBJECTIVE: To assess the outcome of a thalassemia screening program at community hospitals by determining the proportion of at-risk couples able to obtain a prenatal diagnosis (PND) in relation to gestational age (GA). METHODS: We accessed records documenting prenatal screening for thalassemia in lower northern Thailand between January 2014 and December 2016. The proportion of at-risk pregnancies able to obtain a PND was determined and median GAs at the time of at-risk notification were compared. Reasons for failures to obtain PNDs were analyzed. RESULTS: Among 4633 screen-positive couples, 259 (5.6%) were identified as at-risk while 23 were excluded due to unconfirmed outcomes. Forty-one declined a PND and were excluded from the final calculations. Of the 195 remaining couples, 140 (71.8%) obtained a PND. Their median GA at the time of at-risk notification was 12.4 (5.6-29.1) weeks, which was earlier than the median GA of 17.7 (6.9-34.6) weeks for couples not undergoing PND (P < .001). Risks for various types of thalassemia and GA were associated with the chances of achieving a PND. CONCLUSION: In practice, one quarter of couples identified as at-risk were unable to obtain a PND. Time-influencing factors seem to be a major determinant.

Evaluation of a cryopreservation procedure to set up a new bone marrow transplant unit using lymphocyte proliferation test.

A bone marrow transplant (BMT) is one kind of standard treatment modality in advanced hemato-oncology. In order to set up a BMT unit, one of the important steps before starting a clinical program is to evaluate the cryopreservation procedure for stem cell storage. Twenty one bags of buffy coat were used to be the testing specimens. They were processed and frozen according to cryopreservation protocol and kept in liquid nitrogen for 2 weeks. The evaluation process was carried out with a lymphocyte proliferation test together with trypan blue staining. By measuring the optical density of each lymphocyte containing well after stimulation, the lymphocyte proliferation value (LPV) could be obtained. When comparing them before and after cryopreservation, the LPV was 2.064 ± 0.379 (mean ± SD) and 1.913 ± 0.546, (p = 0.314), respectively. At 2 weeks after cryopreservation, comparing between the frozen group and the unfrozen control, the LPV was 1.913 ± 0.546 and 0.486 ± 0.453, (p < 0.05), respectively. The LPV showed clear efficacy of the procedure, especially for preserving the cellular proliferation function. Our model of the cryopreservation procedure evaluation at pre-clinical phase by use of a buffy coat and lymphocyte proliferation test seems feasible for newly-established small BMT units. With these results, clinical transplantations can be performed with more confidence.

Q Sepharose micro-column chromatography: A simple screening method for identifying beta thalassemia traits and hemoglobin E carriers.

OBJECTIVES: For beta thalassemia control program in pregnancy, mass screening of the carrier state by determination of the hemoglobin (Hb) A2 and Hb E proportions and mutation analysis is a preferred method for making prenatal diagnoses. Q Sepharose micro-column chromatography, developed for the determination of Hb A2 and Hb E for screening purposes, was compared with high performance liquid chromatography (HPLC) to ascertain its relative accuracy and reliability. DESIGN AND METHODS: Results using Q Sepharose micro-column chromatography in 350 blood specimens, including 50 samples genetically proven to be beta thalassemia heterozygotes, were compared to HPLC for validation. An additional study was conducted to test a clinical application on a large-scale survey for beta thalassemia in 1581 pregnant women and their spouses. RESULTS: The mean (±SD) Hb A2 proportions in the normal and genetically proven beta thalassemia heterozygotes were 2.70±0.40% and 6.30±1.23%, respectively, as determined by Q-Sepharose micro-column chromatography, and 2.65±0.31% and 5.37±0.96%, respectively, as determined by HPLC. The mean Hb E proportions in the Hb E heterozygotes were 23.25±4.13% and 24.72±3.5% as determined by Q Sepharose micro-column chromatography and HPLC, respectively. In the large-scale survey for beta thalassemia, 23 at risk couples were detected. Seven affected fetuses were identified by prenatal diagnosis. CONCLUSIONS: Q Sepharose micro-column chromatography was found to be reliable, reproducible and well-suited for large-scale surveys. Additionally, by being reusable and convenient, this simple and economical chromatography method may be an alternative means to screen for beta thalassemia and Hb E carriers in the mass population.