Magnetic resonance imaging assessment of the changes of cardiac and hepatic iron load in thalassemia patients before and after hematopoietic stem cell transplantation

To investigate the value of T2* technique on 3.0 T magnetic resonance imaging (MRI) in evaluating the changes of cardiac and hepatic iron load before and after hematopoietic stem cell transplantation (HSCT) in patients with thalassemia (TM), the 141 TM patients were divided into 6 group for subgroup analysis: 6, 12, 18, 24 and > 24 months group, according to the postoperative interval. The T2* values of heart and liver (H-T2*, L-T2*) were quantified in TM patients before and after HSCT using 3.0 T MRI T2* technology, and the corresponding serum ferritin (SF) was collected at the same time, and the changes of the three before and after HSCT were compared. The overall H-T2* (P = 0.001) and L-T2* (P = 0.041) of patients after HSCT were higher than those before HSCT (mean relative changes = 19.63%, 7.19%). The H-T2* (P < 0.001) and L-T2* (P < 0.001) > 24 months after HSCT were significantly higher than those before HSCT (mean relative changes = 69.19%, 93.73%). The SF of 6 months (P < 0.001), 12 months (P = 0.008), 18 months (P = 0.002) and > 24 months (P = 0.001) were significantly higher than those before HSCT (mean relative changes = 57.93%, 73.84%, 128.51%, 85.47%). There was no significant improvement in cardiac and liver iron content in TM patients within 24 months after HSCT, while the reduction of cardiac and liver iron content in patients is obvious when > 24 months after HSCT.


Inclusion and exclusion criteria
Inclusion criteria: (1) patients diagnosed with TM by genetic diagnosis.(2) Any type of HSCT, including bone marrow transplantation (BMT), peripheral blood cell transplantation (PBCT), or (umbilical cord blood transplantation (UCBT).Cardiac and liver GRE sequence MRI examinations were performed before and after HSCT.

Statistical analysis
SPSS 26.0 statistical software package was used to analyze the total H-T 2 * , L-T 2 * , and SF before and after HSCT.The patients were divided into 6-month group (< 6 months), 12-month group (6-12 months), 18-month group (12-18 months), 24-month group (18-24 months) and > 24 months group according to the postoperative interval.Statistical analysis was performed according to the test level α = 0.05.1a shows that the average value of H-T 2 * is 27.17, R 2 (goodness of fit) is 0.9942, and 8 echo SI are included.CMRTools in FIG.1b shows that the average value of L-T 2 * is 1.90, R 2 is 0.999, the SI of the first five echoes are included, and the cross represents the SI of the discarded deviation from the fitted curve.(c,d) β-TM patient, a 14-year-old male, was examined after HSCT.CMRTools in FIG.1c shows that the average value of H-T 2 grading, the number of cases in severe and moderate liver iron concentration after HSCT decreased (preoperative n = 116, 82.27%; Postoperative n = 105, 74.47%), the number of cases in mild and normal groups increased (preoperative n = 25, 17.73%; Postoperative n = 36, 25.53%).After HSCT, the number of patients with severe and moderate cardiac iron concentration decreased (preoperative n = 18, 12.77%; postoperative n = 14, 9.93%), while the number of mild and normal groups increased (preoperative n = 123, 87.23%; postoperative n = 127, 90.07%).Figure 4a shows that H-T 2 * in TM patients increased from 18 months after HSCT (Significant reduction in cardiac iron load); Fig. 4b shows that L-T 2 * in TM patients increased from 24 months after HSCT (Significant reduction in hepatic iron load).

Discussion
The primary treatment for most TM patients is supportive medical care.Patients with severe anemia require regular, lifelong blood transfusion and iron chelation therapy 20 .Advances in HSCT offer potential cure options for some patients.A recent case-control study noted that overall survival was similar in children and adults with TM regardless of HSCT or chronic transfusion therapy (82.6 ± 2.7%, 85.3 ± 2.7%, P = NS) 21 .However, HSCT is associated with improved health-related quality of life compared with chronic transfusion therapy [22][23][24]  www.nature.com/scientificreports/cost of lifelong blood transfusion and conventional treatment with iron chelation therapy 25,26 .However, how does the level of organ iron load change before and after HSCT in TM patients?In this study, the internal organ iron load of TM patients after HSCT was evaluated based on MRI T 2 * technology, and it was found that there was no significant difference in L-T 2 * and H-T 2 * at 24 months after HSCT compared with that before HSCT.For patients more than 24 months after HSCT, L-T 2 * and H-T 2 * were significantly higher than those before HSCT, that is, the iron load of viscera was significantly reduced.This suggests that blood reconstitution in TM patients after HSCT is a slow process, which is the same view as Jagannath 4 .In the process of blood reconstitution, most TM patients still need transfusion therapy.In fact, some of the patients in this study still required regular transfusion therapy after HSCT.Regular blood transfusion is easy to cause iron load.Therefore, it is recommended that TM patients after HSCT should keep regular monitoring of organ iron load in the process of blood reconstitution (at least 24 months after HSCT), and make appropriate iron chelation therapy.
Hematopoietic stem cell transplantation replaces the ineffective erythropoiesis with an effective allogeneic substitute.The transplanted hematopoietic progenitor cells take over the function and synthesise normal red blood cells.The process enables to gradually correct anemia and eliminate the hemolytic process which improves the lifetime need for blood transfusion and chelation therapy 3 .However stem cells enter the bone marrow and, and within 2-4 weeks of transplantation, they begin to produce new white blood cells, red blood cells, and platelets 3,27 .
Blood smear of the first patient with TM undergoing HSCT showed typical changes in TM 27 , initial hemoglobin 5.9 g/dl, white ocyte count 3 × 10 9 /l, 50% nucleated red blood cells.Hemoglobin electrophoresis shows only the fetal haemoglobin.The hematocrit on days 29, 47, 97 day was 37%, 32%, 37%, respectively; white blood cell count was 0.66 × 10 9 /l, 3 × 10 9 /1, 3.4 × 10 9 /l, respectively.Liver and spleen enlargement completely disappeared by day 60.During the sixth month, the patient was well with a hemoglobin of 14.4 g/dl, white blood cell count 3.9 × 10 9 /l, a and fetal haemoglobin undetectable by electrophoresis.The whole process embodies the changing process of blood cells after HSCT in TM patients after HSCT, that is, it seems to take a longer time to show efficacy.
A retrospective analysis of 516 pediatric and adult thalassemia patients suggested that the 30-year survival rate after HSCT was similar to that expected for thalassemia patients receiving conventional care (transfusion, etc.), and that the vast majority of HSCT survivors were cured from thalassemia (94.2%) 21.A large retrospective analysis by Baronciani et al. proposed a cure of 80-90% of thalassemia patients in actively addressing multiple post-HSCT factors, including improved pretreatment regimen, improved graft-versus-host disease prevention (GvHD), and more effective antimicrobial, antiviral and antifungal therapy.Overall, HSCT has a high cure rate for treating patients with thalassemia 28 .
Some studies have shown that there is no significant correlation between myocardial iron load and SF and liver iron load [29][30][31][32] .Chaosuwannakit 33 , Ghugre 34 and Kaltwasser 35 also showed that there was no significant correlation between myocardial iron load and SF, while there was a medium-low correlation between liver iron load and SF.The results of this study were similar, the overall L-T 2 * , H-T 2 * , and SF showed no or low correlation before and after HSCT in TM patients.High correlations in some subdivided subgroups are likely due to overfitting or sampling error resulting from the abrupt reduction in sample size after subgroup analysis.Obviously, the use of SF alone to monitor iron load in humans is not comprehensive, as this measure is likely to show considerable changes due to blood transfusion, inflammation, infection, or other chronic diseases.Some studies have also pointed out that SF may be too low when patients have severe iron deposition 34,36 .In conclusion, these results underscore the importance of T 2 * MRI in assessing iron burden, especially in the liver and heart.In this study, the statistics of red blood cell transfusion and iron removal therapy before HSCT in 115 TM patients were collected, and it was found that the red blood cell transfusion required by TM patients with different disease states was slightly different, 1.5-3 units of red blood cell transfusion accounted for a large proportion.All patients received iron chelation therapy with different methods, but because of the different medical compliance of patients in this region, there were 15 patients who did not receive regular iron chelation therapy.Different iron chelation regimens may affect the iron load of organs.However, more than 90% of the patients received regular iron chelation therapy, and we conducted subgroup analysis at different time points, and the results were reliable.
Limitations of this study: (1) Although 141 TM patients was included, the overall sample size was relatively adequate.However, when analyzing different subgroups, the sample size was relatively small.(2) As most of the TM patients in our center are β-TM, the number of α-TM patients included in our study is small, but it has little impact on the purpose of this study.(3) Due to the limited clinical data we collected, blood routine and liver and kidney function indexes were only presented as baseline data, without considering the main factors that may affect serum ferritin level.(4) This study only explored the changes of heart and liver iron load in TM patients before and after HSCT from the overall change trend, and did not control for the variable of iron chelators, that is, it can not prove that iron status/Mean change rates (%) are not touched by iron chelators, their doses, and amounts of blood transfusions.This point may become an entry point for the future work.
In conclusion, SF alone cannot effectively assess the change of iron load after HSCT.MRI T 2 * technique is of great significance for quantitative assessment of cardiac and hepatic iron content in TM patients after HSCT.At least 24 months after HSCT, the heart and liver iron content of TM patients should be continuously and regularly monitored by SF and MRI, so as to better formulate the corresponding chelating iron removal plan.

( 3 )
Treatment by regular or irregular blood transfusion.(4) Regular or irregular iron removal treatment.(5) Age ≥ 4 years.The exclusion criteria: (1) GRE image data artifacts were large and did not meet the measurement requirements.(2) Patients combined with other chronic liver diseases or tumor diseases.(3) Failure of HSCT or postoperative complications.

Figure 1 .
Figure 1.(a,b) β-TM patient, 25-year-old male, preoperative examination of HSCT.CMRTools in FIG.1ashows that the average value of H-T 2 * is 27.17, R 2 (goodness of fit) is 0.9942, and 8 echo SI are included.CMRTools in FIG.1bshows that the average value of L-T 2 * is 1.90, R 2 is 0.999, the SI of the first five echoes are included, and the cross represents the SI of the discarded deviation from the fitted curve.(c,d) β-TM patient, a 14-year-old male, was examined after HSCT.CMRTools in FIG.1cshows that the average value of H-T 2 Mean change rates of Heart T2* Mean change rates of Liver T2* Mean change rates of Serum FerriƟn

Figure 2 .Figure 3 .
Figure 2. A line plot based on the mean change rates (%) over time of the change in Heart T 2 * (ms), Liver T 2 *

Figure 4 .
Figure 4. Bar graphs of heart T 2 * , liver T 2 * and serum ferritin based on means.

Figure 5 .
Figure 5. Scatter plot (a-c) shows that the fitting degree of trend line and 95% confidence interval (CI) of total liver T 2 * , heart T 2 * and serum ferritin before and after HSCT is very low, showing low correlation or no correlation.