Comparative study of hemoglobin changes following transfusion of fresh and old blood in patients with thalassemia referred to Bahrami Hospital

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Hemoglobin (Hb) after transfusion determines the need for next transfusion. There is controversy regarding the effect of fresh packed RBC compared to old packed RBC transfusion on Hb. This study compared hemoglobin changes following transfusion between fresh and old packed RBC in transfusion-related thalassemia (TRT) patients. Materials and methods In this cohort study, 140 TRT patients (age > 5years) referred to Bahrami Children's Hospital in mid-2013 were included. Equal number (n = 70) of age and gender matched TRT patients with exposure to fresh (stored for less than 7 days) and old (stored for more than 7 days) packed RBC transfusion were followed for 14 days. In both groups, demographic information (age, sex) were recorded at enrolment. Physical examination data and blood parameters, including red blood cells (RBC), white blood cells (WBC), platelets (PLT), Hb, ferritin, transferrin, total iron binding capacity and bilirubin at baseline and follow up. Results Time-group interaction was significant only for WBC (p < 0.001) and Hb (p = 0.029) indicating that fresh packed RBC transfusion resulted in higher Hb and lower WBC at follow up compared to baseline levels. Among the baseline variables only fresh packed RBC transfusion was found to increase the odds of having Hb of at least 1 mg/dL higher than baseline (OR = 15.465, p < 0.001). Conclusions Fresh packed RBC transfusion might increase the interval between transfusions due to more Hb elevation rate after transfusion in thalassemia major. Hematological factors stored blood old blood fresh blood children thalassemia Figures Figure 1 Introduction Thalassemia is the most common genetic hematologic disorder that manifests with anemia caused by defect in the production of hemoglobin (Hb) ( 1 ). Beta thalassemia is caused by reduced production of beta globin chain in Hb, can affect various organs, and may result in morbidity and mortality ( 2 ). Alpha thalassemia is among the common genetic defects in Hb production and is caused by lack or reduces synthesis of alpha globin chain ( 3 , 4 ). The global prevalence of thalassemia is reported to range between 0.2 and 27.2 per 100000 with alpha and beta thalassemia prevalence ranging from 0.03–4.5 and 0.2–49.6 per 100000, respectively ( 5 ). The prevalence of thalassemia is higher in regions called thalassemia belt, including Mediterranean region, North and Eastern Africa, Middle-East, Indian sub-continent, South-East Asia, and South America ( 6 , 7 ). In Iran, the prevalence of thalassemia is 11.35% (range 8.5% to 14.92%), which has not changed significantly in the past decade ( 8 ). Thalassemia symptoms can range from asymptomatic patients to severe anemia, and hepatosplenomegaly that might result in death if not treated ( 9 ). Due to the wide range of complications, thalassemia is considered among the chronic diseases of interest in Iran ( 10 ). Thalassemia major is defined as severe anemia requiring blood transfusion, ineffective erythropoiesis, and intramedullary hematopoiesis ( 11 ). Blood transfusion is the standard treatment of Beta thalassemia major and is recommended to maintain Hb levels between 9.5 and 10.5 mg/dL ( 4 ). Although lifesaving, chronic blood transfusion can result in complications that may also affect survival in patients with thalassemia ( 12 ). The most common complications of blood transfusion in thalassemia are iron overload, alloimmunization against Rh antigens, and splenomegaly ( 12 , 13 ). Iron overload can damage other organs including liver, heart, pancreas and kidneys ( 14 ). The decrease in Hb level after blood transfusion dictates the time for next transfusion (two to five weeks) ( 15 ). Different factors may affect the Hb reduction rate after blood transfusion ( 16 ). Hypersplenism and splenomegaly affects Hb levels after transfusion making it necessary to perform splenectomy to increase Hb levels and elongate the between transfusion intervals ( 3 ). Another factor is the quality of transfused packed RBC ( 16 ). RBC can be stored for 42 days before transfusion ( 17 ). Blood cells undergo a series of physiological changes during storage ( 18 ). These changes are called storage lesions and include Hb oxidation, Hb release from red blood cells (RBCs), structural breakdown of RBC membrane, and increased cell breakdown ( 19 ). Furthermore, metabolic changes, including lactate and 2,3 di-phosphoglycerate accumulation, and reduces adenosine three phosphate (ATP) can also happen during storage ( 20 ). Biochemical evidence indicate that old RBCs are at increased risk for lysis and inducing inflammation ( 21 ). While these complications might be well tolerated in a healthy individual with stable hemodynamic condition, they may compromise unstable patients ( 19 ). Therefore, the provision of a high quality packed RBC to transfusion-dependent patients with thalassemia could increase transfusion intervals and reduce complications and improve quality of life and survival in these patients ( 22 ). Therefore, it is hypothesized that old blood transfusion may result in hemolysis and releasing Hb and iron and increasing inflammation ( 23 ). There is no solid definition for fresh pack RBC; however, in most trials, fresh packed RBC is defined as packed RBC stored for less than 14 days or sometimes less than the median storage duration in each center ( 24 , 25 ). While some studies indicated increased mortality, extravascular hemolysis, increased serum saturated and total transferrin level, as well as free iron and E-Coli infection in patients receiving old packed RBC transfusion (2–6 weeks) compared to fresh packed RBC transfusion (less than 10 days) ( 19 , 26 – 28 ), other studies indicated no significant difference in mortality, multiple organ dysfunction ( 29 , 30 ). These challenges have resulted in reducing the recommended packed RBC storage duration to less than 5 weeks (21 to 35 days) in some recommendations ( 31 ). However, there is no conclusive evidence on the comparative effect of old and fresh packed RBC transfusion on hematological factors in patients with thalassemia. Therefore, the aim of this study was to compare changes in hematologic factors between fresh (24 hour) and old (more than 7 days) packed RBC transfusion in transfusion-dependent patients with thalassemia. Methods Study design This cohort study was conducted on patients with thalassemia who referred to the Bahrami Children’s Hospital in mid-2013. The inclusion criteria were transfusion-dependent thalassemia, age older than 5, and willingness to participate in the study. Exclusion criteria with any accompanying conditions or indications for transfusion other than thalassemia, including trauma and surgery. Sample size Sample size was calculated based on mean difference equation considering type I and II errors of 0.05 and 0.2, respectively. $$n=\frac{{{(Z}_{1-\raisebox{1ex}{$\alpha$}\!\left/\!\raisebox{-1ex}{$2$}\right.}+{Z}_{1-\beta})}^{2}({\delta}_{1}^{2}+{\delta}_{2}^{2})}{{({\mu}_{1}-{\mu}_{2})}^{2}}$$ The final sample size was 70 patients in each group (140 patients in total). Study procedure Researchers approached eligible patients and a written informed consent was obtained from willing patients after providing a brief information about the study. Primarily blood group was identified for all patients in order to rule out alloimmunization. Transfusion blood was collected from donors based on the standard protocols in transfusion bags. Screening was performed for transfusion transmitted infections (TTIs), including hepatitis B, hepatitis C, and Syphilis. Then, transfusion bags were centrifuged at 4000 RPM to separate plasma. Separated plasma was stored in a separate bag and the RCC was stored in transfusion bank. Fresh packed RBC were transfused within 48 hours after donation while old packed RBC were transfused within 7 days from donation ( 32 ). Demographic data (age and gender) and physical examination were collected from both the fresh and old blood groups. Before transfusion, blood samples were obtained from all patients to assess red blood cell (RBC) count, white blood cell (WBC) count, Hb, transferrin, total iron binding capacity (TIBC), serum ferritin, and bilirubin. Follow up assessment was performed four weeks after transfusion and included blood sampling and physical examination. Infection was defined as WBC count above 10000 /dL. Statistical analysis The statistical package for social sciences (SPSS) software version 22 (IBM Inc, USA) was used to analyze data. Normality of data was assessed using the Kolmogorov-Smirnov test. Mean and standard deviation (SD) were used to present continuous data. Categorical variables were shown using frequency and percentage. Comparison of the categorical variables between groups was performed using the chi-square or Fisher exact tests. Number need to treat (NNT) was calculated for Hb using the following equation; $$NNT=\frac{1}{\left|Controleventrate-Experimentaleventrate\right|}$$ The repeated measures analysis of variance (ANOVA) analysis was used to evaluate the effect of time, group, and time-group interaction on hematological and biochemical parameters. Binary logistic regression was used to identify the baseline predictors for at least 1 mg/dL higher Hb at follow up compared to baseline values by considering Hb change as dependent variable and demographic and baseline hematologic and biochemical parameters as dependent variables. The level of statistical significance was p < 0.05. Results Of the 140 patients in the study, 70 received fresh packed RBC and 70 received old packed RBC transfusion. Comparison of baseline data between groups is shown in Table 1 . There was no significant difference between groups in terms of demographic, hematologic, and Biochemical parameters at baseline (p > 0.05). Table 1 Comparison of demographic parameters between fresh and old packed RBC transfusion patients Variable Fresh packed RBC Old packed RBC p Demographic Age (year) 19.00 ± 2.32 19.17 ± 2.79 0.693† Gender Female 39 (55.71%) 31 (44.28%) 0.237‡ Male 31 (44.28%) 39 (55.71%) WBC: white blood cell, Hb: hemoglobin, PLT: platelet † Independent t-test was used for the comparison ‡ The chi-square test was used for the comparison Changes in hematologic and biochemical parameters from baseline to follow up are shown in Table 2 . Based on the repeated measures ANOVA analysis, there was a significant time effect for Hb (p < 0.001), PLT (p,0.001), and TIBC (p < 0.001) indicating a significant change over time regardless of the type of transfusion. There was a significant group effect for PLT (p = 0.010) indicating that type of transfusion resulted in a different change in PLT. However, time-group interaction was significant only for WBC (p < 0.001) and Hb (p = 0.029) indicating a different trend in these two variables over time between groups (Fig. 1 ). Table 2 Changes in hematologic, and biochemical parameters from baseline to follow up among fresh and old packed RBC transfusion patients Variable Fresh packed RBC Old packed RBC Time effect p Group effect p Time-group interaction p Baseline Follow up Baseline Follow up Hematologic WBC (/dL) 7724.28 ± 2663.73 7093.11 ± 2064.75 7662.85 ± 2130.67 8052.57 ± 2224.67 0.064 0.240 < 0.001* Hb (mg/dL) 9.31 ± 0.97 9.96 ± 1.00 9.00 ± 0.98 9.41 ± 0.96 < 0.001* 0.007 0.029* PLT (/dL) 428785.71 ± 207273.94 384585.71 ± 139676.17 455571.42 ± 204564.33 428857.14 ± 174339.43 < 0.001* 0.010* 0.257 Biochemical Ferritin (ng/mL) 2487.71 ± 1546.78 2460.40 ± 1593.20 2736.02 ± 1478.60 2624.91 ± 1590.63 0.341 0.415 0.564 TIBC (µg/dL) 218.22 ± 36.79 234.40 ± 47.36 223.71 ± 41.41 242.58 ± 49.76 < 0.001* 0.302 0.698 Total bilirubin (mg/dL) 2.69 ± 1.96 2.44 ± 2.02 2.94 ± 1.94 2.34 ± 1.77 0.083 0.724 0.479 WBC: white blood cell, Hb: hemoglobin, PLT: platelet, TIBC: total iron binding capacity Repeated measures analysis of variance was used for the analysis * Significant effect at α = 0.05 At the fourth week follow up Hb was at least 1 mg/dL higher than baseline in 21 (30%) of patients in the fresh packed RBC transfusion compared to 2 (2.9%) in the old packed RBC transfusion group (p < 0.001). Therefore, the NNT was 5.85. Logistic regression analysis was perform to identify the baseline parameters that could predict elevated Hb level at the forth week follow up (Table 3 ). There was only a significant relationship between transfusion group and higher Hb at follow up (OR = 15.465, 95% CI for OR: 3.328–71.865, p < 0.001) indicating that compared to old packed RBC transfusion, fresh packed RBD transfusion was associated with 15.465 times increased odds of having at least 1 mg/dL higher Hb level at follow up. Table 3 Relationship between at least 1 mg/dL higher hemoglobin at follow up and demographic and baseline parameters Variable p OR 95% CI for OR Lower Upper Group (Fresh packed RBC transfusion) < 0.001* 15.465 3.328 71.865 TIBC 0.216 0.989 0.971 1.007 Gender (Female) 0.920 0.949 0.338 2.665 Age 0.514 0.929 0.745 0.1.159 WBC 0.548 1.000 1.000 1.000 Hb 0.812 0.923 0.473 1.783 PLT 0.639 1.000 1.000 1.000 Ferritin 0.725 1.000 1.000 1.000 Total bilirubin 0.680 0.940 0.698 1.264 WBC: white blood cell, Hb: hemoglobin, PLT: platelet, TIBC: total iron binding capacity Binary logistic regression was used for the analysis with less than 1 mg/dL increase in Hb at follow up as reference dependent variable and old packed RBC transfusion and male gender were set as reference independent variables. * Significant relationship at α = 0.05 Discussion The findings of this study showed that receiving fresh packed RBC was associated with higher Hb at follow up compared to old packed RBC transfusion. The percentage of patients with at least 1 mg/dL higher Hb compared to baseline was significantly higher among patients in the fresh packed RBC transfused compared to the old packed RBC transfused group. This study also indicated that WBC and PLT reduced to a more extent at follow up in the fresh packed PRBC transfused compared to the old packed RBC transfused group. These findings may indicate that fresh packed RBC transfusion might have resulted in slower Hb reduction due to less inflammation. Storage can affect RBCs in through oxidative damage to Hb, RBC membrane, cell proteins, and metabolic impairments, including the accumulation of lactate and reduced level of 2,3 di-phosphoglycerate in RBC ( 33 ). Oxidative damage to RBC can result in the separation of Haeme from Hb and damage to structural proteins and lipids that impair deformability and metabolism of stored RBCs through the production of inflammatory cytokines and reduced nitric oxide capacity ( 34 ). Therefore, transfused RBCs might adhere to the endothelium and activate inflammation in the recipient. Although these changes were found to be well tolerated in healthy recipients, these changes might have a significant effect in critically ill patients and patients with hematologic abnormalities including thalassemia ( 35 – 37 ). This study showed that receiving fresh packed RBC resulted in increased odds of having at least 1 mg/dL higher Hb in follow up compared to the baseline Hb. There is controversy in the effectiveness of fresh packed RBC transfusion in achieving a higher Hb level after transfusion. While some studies indicated a significantly higher Hb after transfusion of fresh packed RBC compared to old packed RBC ( 38 , 39 ), some studies did not find any difference in Hb change between fresh and old packed RBC transfusion ( 19 , 33 , 40 ). The reason for these differences could be attributed to the differences in sample size, presence of hepatosplenomegaly, splenectomy, and having critical illness as well as differences in the definition of fresh and old packed RBC transfusion in some studies. Furthermore, the findings of this study indicated that higher proportion of fresh packed RBC transfused patients had post transfusion Hb at least 1 mg/dL higher than baseline levels compared to old packed RBC transfused patients. In other words, in order to have one patient with Hb of at least 1 mg/dL higher than baseline, six patients should receive fresh packed RBC transfusion (NNT = 5.85). These findings indicate that using fresh packed RBC increases the odds of having higher levels of Hb to the extent that could prolong next transfusion for at least one week. The findings of this study indicated that fresh packed RBC transfusion resulted in a significant time-group interaction effect on WBC indicating that fresh packed RBC transfusion resulted in lower WBC compared to old packed RBC transfusion. This finding was in line with the findings of previous studies and might be indicative of the activation of host immune response due to packed RBC transfusion mainly through inflammation ( 19 , 41 , 42 ). The hypothesized reason for this finding is the leukocytosis due to the increased cytokines line interleukin-6 (IL-6) and IL-8 in response to high concentration of non-transferrin bound iron in RBC ( 41 ). Therefore, the observed higher WBC in the old packed RBC transfused patients in this study might indicate the higher level of inflammation in these patients compared to those who received fresh packed RBC. The overall findings of this study were in favor of using fresh packed RBC transfusion in patients with thalassemia major in order to achieve higher Hb levels and prolong transfusion intervals. These findings were in line with the current recommendations in Japan and the United Kingdom that prioritize the use fresher packed RBC transfusion ( 31 ). Although cost effectiveness was not assessed in this study, It can be hypothesized that the use of fresh packed RBC might reduce the health and economic burden of thalassemia major by reducing the number of packed RBC transfusion in long term. In Iran the estimated number of patients with beta thalassemia major is 25000 with median estimated life expectancy of 57 years ( 43 , 44 ). Considering the 3-5-week transfusion interval and the required two packed RBC bags in each transfusion, delaying transfusion for one week may result in the saving of packed RBC units over time. It was previously shown that 20–60% of the economic burden of beta thalassemia was related to packed RBC transfusion costs ( 45 – 47 ). In a study conducted in Iran in 2018, the yearly cost of beta thalassemia major was 8321.8 United States Dollar ( 48 ). High packed RBC transfusion frequency was found to attribute to higher serum ferritin and chelation therapy and thus increased economic burden of disease in a multicenter study in Thailand ( 49 ). Besides the direct costs of packed RBC transfusion, including cross matching, leukoreduction, antibody evaluation, blood group and Rh matching, indirect costs, including the cost of transportation also attribute to the economic burden of thalassemia major ( 46 , 49 ). Therefore, considering the effect of using fresh packed RBC can have a long term financial and health benefits for patients with beta thalassemia major through reducing the transfusion frequency. This study had some limitations. One of the limitations of this study was the lack of long-term follow up of the patients. It would be more important to document the long-term effects of receiving fresh packed RBC compared to old packed RBC in patients with thalassemia major. Another limitation of this study was lack of evaluating inflammatory markers due to financial limitations; therefore, it is recommended that further studies evaluate inflammatory cytokines in relation to Hb levels and transfusion intervals in patients with thalassemia major. Conclusion This study reported that fresh packed RBC transfusion might delay the time of next transfusion due to the higher hemoglobin than baseline four weeks after transfusion possibly due to lower inflammatory properties in fresh packed RBC compared to old packed RBC. Abbreviations ANOVA Analysis of Variance Hb Hemoglobin IL-6 Interleukin 6 NNT Number Need to Treat PLT Platelet RBC Red Blood Cell TIBC Total Iron-Binding Capacity TTI Transfusion Transmitted Infection WBC White Blood Cell Declarations Ethics approval and consent to participate The study was conducted in accordance with the Helsinki Declaration and was approved by the Ethics Committee of the Tehran University of Medical Sciences (Ethics code: IR.TUMS.MEDICINE.REC.1403.035). Written informed consent was obtained from all patients or their guardians prior to participating in the study. All personal identifiers were deleted from the collected data to ensure anonymity. Consent for publication All co-authors read and approved the final version of the manuscript and consented for publication. Availability of data and materials The datasets used and analyzed during the current study are available from the corresponding author on reasonable request. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8928118","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":603846311,"identity":"b2afb2c7-d428-4706-b3c2-4004e17c18e5","order_by":0,"name":"Mitra Ardakani Moghadm","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Mitra","middleName":"Ardakani","lastName":"Moghadm","suffix":""},{"id":603846312,"identity":"93d99dd2-880b-4d7e-8320-23468805aac5","order_by":1,"name":"zeynab salami","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"zeynab","middleName":"","lastName":"salami","suffix":""},{"id":603846313,"identity":"d76d31bb-a824-46ca-9c1d-236b1f66d06a","order_by":2,"name":"Mohammad kaji yazdi","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA+UlEQVRIiWNgGAWjYDACHgYGgwQQ4zD/gwMfgDQbO/FaeBgfzgBpYSZCCwQc4GE2BnMIadHtOfyg4GGbnT3fcd5j0ja/tsnzMTMwfviYg1uL2dk2A4PEtmRmycN8adK5fbcN25gZmCVnbsOj5TwDSAszm8FhBjPp3J7bjEAtbMy8eLWwfwBqqecBa7HsuW1PWMvZHpAthyUMDvMYGzP8uJ1IWMuZMwUGCeeOG0geZkt82NtwO7mNmbEZv1/OpG8z/FFWbc93/vCBAz/+3Lad39588MNHPFqAgM2AkQ3KZGwDkw141QMB8wOGPzD2H3wKR8EoGAWjYKQCAEOMUkTcNLYWAAAAAElFTkSuQmCC","orcid":"","institution":"","correspondingAuthor":true,"prefix":"","firstName":"Mohammad","middleName":"kaji","lastName":"yazdi","suffix":""}],"badges":[],"createdAt":"2026-02-20 17:38:11","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8928118/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8928118/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":104593665,"identity":"629048bb-e809-46b4-af44-7ecd656cb9f4","added_by":"auto","created_at":"2026-03-13 17:42:34","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":54274,"visible":true,"origin":"","legend":"\u003cp\u003eChanges in Hemoglobin (A) and white blood cell count (B) from baseline to follow up as per fresh packed RBC and old packed RBC transfusion groups\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8928118/v1/9695fbbad842aa340c0a1f7e.jpeg"},{"id":104781838,"identity":"3ba8bd6b-8f71-40b0-8be5-613958ba8b9f","added_by":"auto","created_at":"2026-03-17 07:56:25","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":723617,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8928118/v1/d65f3830-4888-4751-9845-b02d00654d80.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Comparative study of hemoglobin changes following transfusion of fresh and old blood in patients with thalassemia referred to Bahrami Hospital","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThalassemia is the most common genetic hematologic disorder that manifests with anemia caused by defect in the production of hemoglobin (Hb) (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e). Beta thalassemia is caused by reduced production of beta globin chain in Hb, can affect various organs, and may result in morbidity and mortality (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e). Alpha thalassemia is among the common genetic defects in Hb production and is caused by lack or reduces synthesis of alpha globin chain (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). The global prevalence of thalassemia is reported to range between 0.2 and 27.2 per 100000 with alpha and beta thalassemia prevalence ranging from 0.03\u0026ndash;4.5 and 0.2\u0026ndash;49.6 per 100000, respectively (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e). The prevalence of thalassemia is higher in regions called thalassemia belt, including Mediterranean region, North and Eastern Africa, Middle-East, Indian sub-continent, South-East Asia, and South America (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e). In Iran, the prevalence of thalassemia is 11.35% (range 8.5% to 14.92%), which has not changed significantly in the past decade (\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e). Thalassemia symptoms can range from asymptomatic patients to severe anemia, and hepatosplenomegaly that might result in death if not treated (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e). Due to the wide range of complications, thalassemia is considered among the chronic diseases of interest in Iran (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e). Thalassemia major is defined as severe anemia requiring blood transfusion, ineffective erythropoiesis, and intramedullary hematopoiesis (\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e). Blood transfusion is the standard treatment of Beta thalassemia major and is recommended to maintain Hb levels between 9.5 and 10.5 mg/dL (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). Although lifesaving, chronic blood transfusion can result in complications that may also affect survival in patients with thalassemia (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e). The most common complications of blood transfusion in thalassemia are iron overload, alloimmunization against Rh antigens, and splenomegaly (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIron overload can damage other organs including liver, heart, pancreas and kidneys (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e). The decrease in Hb level after blood transfusion dictates the time for next transfusion (two to five weeks) (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e). Different factors may affect the Hb reduction rate after blood transfusion (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e). Hypersplenism and splenomegaly affects Hb levels after transfusion making it necessary to perform splenectomy to increase Hb levels and elongate the between transfusion intervals (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). Another factor is the quality of transfused packed RBC (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e). RBC can be stored for 42 days before transfusion (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e). Blood cells undergo a series of physiological changes during storage (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e). These changes are called storage lesions and include Hb oxidation, Hb release from red blood cells (RBCs), structural breakdown of RBC membrane, and increased cell breakdown (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e). Furthermore, metabolic changes, including lactate and 2,3 di-phosphoglycerate accumulation, and reduces adenosine three phosphate (ATP) can also happen during storage (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e). Biochemical evidence indicate that old RBCs are at increased risk for lysis and inducing inflammation (\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e). While these complications might be well tolerated in a healthy individual with stable hemodynamic condition, they may compromise unstable patients (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e). Therefore, the provision of a high quality packed RBC to transfusion-dependent patients with thalassemia could increase transfusion intervals and reduce complications and improve quality of life and survival in these patients (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e). Therefore, it is hypothesized that old blood transfusion may result in hemolysis and releasing Hb and iron and increasing inflammation (\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThere is no solid definition for fresh pack RBC; however, in most trials, fresh packed RBC is defined as packed RBC stored for less than 14 days or sometimes less than the median storage duration in each center (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e). While some studies indicated increased mortality, extravascular hemolysis, increased serum saturated and total transferrin level, as well as free iron and E-Coli infection in patients receiving old packed RBC transfusion (2\u0026ndash;6 weeks) compared to fresh packed RBC transfusion (less than 10 days) (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan additionalcitationids=\"CR27\" citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e), other studies indicated no significant difference in mortality, multiple organ dysfunction (\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e). These challenges have resulted in reducing the recommended packed RBC storage duration to less than 5 weeks (21 to 35 days) in some recommendations (\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e). However, there is no conclusive evidence on the comparative effect of old and fresh packed RBC transfusion on hematological factors in patients with thalassemia. Therefore, the aim of this study was to compare changes in hematologic factors between fresh (24 hour) and old (more than 7 days) packed RBC transfusion in transfusion-dependent patients with thalassemia.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStudy design\u003c/h2\u003e \u003cp\u003eThis cohort study was conducted on patients with thalassemia who referred to the Bahrami Children\u0026rsquo;s Hospital in mid-2013. The inclusion criteria were transfusion-dependent thalassemia, age older than 5, and willingness to participate in the study. Exclusion criteria with any accompanying conditions or indications for transfusion other than thalassemia, including trauma and surgery.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eSample size\u003c/h3\u003e\n\u003cp\u003eSample size was calculated based on mean difference equation considering type I and II errors of 0.05 and 0.2, respectively.\u003cdiv id=\"Equa\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equa\" name=\"EquationSource\"\u003e\n$$n=\\frac{{{(Z}_{1-\\raisebox{1ex}{$\\alpha$}\\!\\left/\\!\\raisebox{-1ex}{$2$}\\right.}+{Z}_{1-\\beta})}^{2}({\\delta}_{1}^{2}+{\\delta}_{2}^{2})}{{({\\mu}_{1}-{\\mu}_{2})}^{2}}$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eThe final sample size was 70 patients in each group (140 patients in total).\u003c/p\u003e\n\u003ch3\u003eStudy procedure\u003c/h3\u003e\n\u003cp\u003e Researchers approached eligible patients and a written informed consent was obtained from willing patients after providing a brief information about the study. Primarily blood group was identified for all patients in order to rule out alloimmunization.\u003c/p\u003e \u003cp\u003eTransfusion blood was collected from donors based on the standard protocols in transfusion bags. Screening was performed for transfusion transmitted infections (TTIs), including hepatitis B, hepatitis C, and Syphilis. Then, transfusion bags were centrifuged at 4000 RPM to separate plasma. Separated plasma was stored in a separate bag and the RCC was stored in transfusion bank. Fresh packed RBC were transfused within 48 hours after donation while old packed RBC were transfused within 7 days from donation (\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eDemographic data (age and gender) and physical examination were collected from both the fresh and old blood groups. Before transfusion, blood samples were obtained from all patients to assess red blood cell (RBC) count, white blood cell (WBC) count, Hb, transferrin, total iron binding capacity (TIBC), serum ferritin, and bilirubin. Follow up assessment was performed four weeks after transfusion and included blood sampling and physical examination. Infection was defined as WBC count above 10000 /dL.\u003c/p\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eStatistical analysis\u003c/h2\u003e \u003cp\u003eThe statistical package for social sciences (SPSS) software version 22 (IBM Inc, USA) was used to analyze data. Normality of data was assessed using the Kolmogorov-Smirnov test. Mean and standard deviation (SD) were used to present continuous data. Categorical variables were shown using frequency and percentage. Comparison of the categorical variables between groups was performed using the chi-square or Fisher exact tests. Number need to treat (NNT) was calculated for Hb using the following equation;\u003cdiv id=\"Equb\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equb\" name=\"EquationSource\"\u003e\n$$NNT=\\frac{1}{\\left|Controleventrate-Experimentaleventrate\\right|}$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eThe repeated measures analysis of variance (ANOVA) analysis was used to evaluate the effect of time, group, and time-group interaction on hematological and biochemical parameters. Binary logistic regression was used to identify the baseline predictors for at least 1 mg/dL higher Hb at follow up compared to baseline values by considering Hb change as dependent variable and demographic and baseline hematologic and biochemical parameters as dependent variables. The level of statistical significance was p\u0026thinsp;\u0026lt;\u0026thinsp;0.05.\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cp\u003eOf the 140 patients in the study, 70 received fresh packed RBC and 70 received old packed RBC transfusion. Comparison of baseline data between groups is shown in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e. There was no significant difference between groups in terms of demographic, hematologic, and Biochemical parameters at baseline (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eComparison of demographic parameters between fresh and old packed RBC transfusion patients\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eVariable\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eFresh packed RBC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eOld packed RBC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003ep\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"5\" nameend=\"c5\" namest=\"c1\"\u003e \u003cp\u003eDemographic\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003eAge (year)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e19.00\u0026thinsp;\u0026plusmn;\u0026thinsp;2.32\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e19.17\u0026thinsp;\u0026plusmn;\u0026thinsp;2.79\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.693\u0026dagger;\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eGender\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eFemale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e39 (55.71%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e31 (44.28%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003e0.237\u0026Dagger;\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eMale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e31 (44.28%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e39 (55.71%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"5\"\u003eWBC: white blood cell, Hb: hemoglobin, PLT: platelet\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e\u0026dagger; Independent t-test was used for the comparison\u003c/p\u003e \u003cp\u003e\u0026Dagger; The chi-square test was used for the comparison\u003c/p\u003e \u003cp\u003eChanges in hematologic and biochemical parameters from baseline to follow up are shown in Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e. Based on the repeated measures ANOVA analysis, there was a significant time effect for Hb (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001), PLT (p,0.001), and TIBC (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001) indicating a significant change over time regardless of the type of transfusion. There was a significant group effect for PLT (p\u0026thinsp;=\u0026thinsp;0.010) indicating that type of transfusion resulted in a different change in PLT. However, time-group interaction was significant only for WBC (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001) and Hb (p\u0026thinsp;=\u0026thinsp;0.029) indicating a different trend in these two variables over time between groups (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eChanges in hematologic, and biochemical parameters from baseline to follow up among fresh and old packed RBC transfusion patients\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"8\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eVariable\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c3\" namest=\"c2\"\u003e \u003cp\u003eFresh packed RBC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003eOld packed RBC\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eTime effect\u003c/p\u003e \u003cp\u003ep\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eGroup effect\u003c/p\u003e \u003cp\u003ep\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eTime-group interaction\u003c/p\u003e \u003cp\u003ep\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eBaseline\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eFollow up\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eBaseline\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eFollow up\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"8\" nameend=\"c8\" namest=\"c1\"\u003e \u003cp\u003eHematologic\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eWBC (/dL)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7724.28\u0026thinsp;\u0026plusmn;\u0026thinsp;2663.73\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e7093.11\u0026thinsp;\u0026plusmn;\u0026thinsp;2064.75\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e7662.85\u0026thinsp;\u0026plusmn;\u0026thinsp;2130.67\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e8052.57\u0026thinsp;\u0026plusmn;\u0026thinsp;2224.67\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.064\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.240\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.001*\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eHb (mg/dL)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e9.31\u0026thinsp;\u0026plusmn;\u0026thinsp;0.97\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9.96\u0026thinsp;\u0026plusmn;\u0026thinsp;1.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e9.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.98\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e9.41\u0026thinsp;\u0026plusmn;\u0026thinsp;0.96\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.001*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.007\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.029*\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePLT (/dL)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e428785.71\u0026thinsp;\u0026plusmn;\u0026thinsp;207273.94\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e384585.71\u0026thinsp;\u0026plusmn;\u0026thinsp;139676.17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e455571.42\u0026thinsp;\u0026plusmn;\u0026thinsp;204564.33\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e428857.14\u0026thinsp;\u0026plusmn;\u0026thinsp;174339.43\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.001*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.010*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.257\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"8\" nameend=\"c8\" namest=\"c1\"\u003e \u003cp\u003eBiochemical\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFerritin (ng/mL)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2487.71\u0026thinsp;\u0026plusmn;\u0026thinsp;1546.78\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2460.40\u0026thinsp;\u0026plusmn;\u0026thinsp;1593.20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2736.02\u0026thinsp;\u0026plusmn;\u0026thinsp;1478.60\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2624.91\u0026thinsp;\u0026plusmn;\u0026thinsp;1590.63\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.341\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.415\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.564\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTIBC (\u0026micro;g/dL)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e218.22\u0026thinsp;\u0026plusmn;\u0026thinsp;36.79\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e234.40\u0026thinsp;\u0026plusmn;\u0026thinsp;47.36\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e223.71\u0026thinsp;\u0026plusmn;\u0026thinsp;41.41\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e242.58\u0026thinsp;\u0026plusmn;\u0026thinsp;49.76\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.001*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.302\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.698\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTotal bilirubin (mg/dL)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.69\u0026thinsp;\u0026plusmn;\u0026thinsp;1.96\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.44\u0026thinsp;\u0026plusmn;\u0026thinsp;2.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2.94\u0026thinsp;\u0026plusmn;\u0026thinsp;1.94\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2.34\u0026thinsp;\u0026plusmn;\u0026thinsp;1.77\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.083\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.724\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.479\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"8\"\u003eWBC: white blood cell, Hb: hemoglobin, PLT: platelet, TIBC: total iron binding capacity\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eRepeated measures analysis of variance was used for the analysis\u003c/p\u003e \u003cp\u003e* Significant effect at α\u0026thinsp;=\u0026thinsp;0.05\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eAt the fourth week follow up Hb was at least 1 mg/dL higher than baseline in 21 (30%) of patients in the fresh packed RBC transfusion compared to 2 (2.9%) in the old packed RBC transfusion group (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001). Therefore, the NNT was 5.85.\u003c/p\u003e \u003cp\u003eLogistic regression analysis was perform to identify the baseline parameters that could predict elevated Hb level at the forth week follow up (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). There was only a significant relationship between transfusion group and higher Hb at follow up (OR\u0026thinsp;=\u0026thinsp;15.465, 95% CI for OR: 3.328\u0026ndash;71.865, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001) indicating that compared to old packed RBC transfusion, fresh packed RBD transfusion was associated with 15.465 times increased odds of having at least 1 mg/dL higher Hb level at follow up.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eRelationship between at least 1 mg/dL higher hemoglobin at follow up and demographic and baseline parameters\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eVariable\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003ep\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eOR\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e95% CI for OR\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eLower\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eUpper\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGroup (Fresh packed RBC transfusion)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.001*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e15.465\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e3.328\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e71.865\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTIBC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.216\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.989\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.971\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.007\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGender (Female)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.920\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.949\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.338\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e2.665\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAge\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.514\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.929\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.745\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.1.159\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eWBC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.548\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eHb\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.812\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.923\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.473\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.783\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.639\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFerritin\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.725\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.000\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTotal bilirubin\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.680\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.940\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.698\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.264\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"5\"\u003eWBC: white blood cell, Hb: hemoglobin, PLT: platelet, TIBC: total iron binding capacity\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eBinary logistic regression was used for the analysis with less than 1 mg/dL increase in Hb at follow up as reference dependent variable and old packed RBC transfusion and male gender were set as reference independent variables.\u003c/p\u003e \u003cp\u003e* Significant relationship at α\u0026thinsp;=\u0026thinsp;0.05\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe findings of this study showed that receiving fresh packed RBC was associated with higher Hb at follow up compared to old packed RBC transfusion. The percentage of patients with at least 1 mg/dL higher Hb compared to baseline was significantly higher among patients in the fresh packed RBC transfused compared to the old packed RBC transfused group. This study also indicated that WBC and PLT reduced to a more extent at follow up in the fresh packed PRBC transfused compared to the old packed RBC transfused group. These findings may indicate that fresh packed RBC transfusion might have resulted in slower Hb reduction due to less inflammation.\u003c/p\u003e \u003cp\u003eStorage can affect RBCs in through oxidative damage to Hb, RBC membrane, cell proteins, and metabolic impairments, including the accumulation of lactate and reduced level of 2,3 di-phosphoglycerate in RBC (\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e). Oxidative damage to RBC can result in the separation of Haeme from Hb and damage to structural proteins and lipids that impair deformability and metabolism of stored RBCs through the production of inflammatory cytokines and reduced nitric oxide capacity (\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e). Therefore, transfused RBCs might adhere to the endothelium and activate inflammation in the recipient. Although these changes were found to be well tolerated in healthy recipients, these changes might have a significant effect in critically ill patients and patients with hematologic abnormalities including thalassemia (\u003cspan additionalcitationids=\"CR36\" citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThis study showed that receiving fresh packed RBC resulted in increased odds of having at least 1 mg/dL higher Hb in follow up compared to the baseline Hb. There is controversy in the effectiveness of fresh packed RBC transfusion in achieving a higher Hb level after transfusion. While some studies indicated a significantly higher Hb after transfusion of fresh packed RBC compared to old packed RBC (\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e, \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e), some studies did not find any difference in Hb change between fresh and old packed RBC transfusion (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e, \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e). The reason for these differences could be attributed to the differences in sample size, presence of hepatosplenomegaly, splenectomy, and having critical illness as well as differences in the definition of fresh and old packed RBC transfusion in some studies. Furthermore, the findings of this study indicated that higher proportion of fresh packed RBC transfused patients had post transfusion Hb at least 1 mg/dL higher than baseline levels compared to old packed RBC transfused patients. In other words, in order to have one patient with Hb of at least 1 mg/dL higher than baseline, six patients should receive fresh packed RBC transfusion (NNT\u0026thinsp;=\u0026thinsp;5.85). These findings indicate that using fresh packed RBC increases the odds of having higher levels of Hb to the extent that could prolong next transfusion for at least one week.\u003c/p\u003e \u003cp\u003eThe findings of this study indicated that fresh packed RBC transfusion resulted in a significant time-group interaction effect on WBC indicating that fresh packed RBC transfusion resulted in lower WBC compared to old packed RBC transfusion. This finding was in line with the findings of previous studies and might be indicative of the activation of host immune response due to packed RBC transfusion mainly through inflammation (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e, \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e). The hypothesized reason for this finding is the leukocytosis due to the increased cytokines line interleukin-6 (IL-6) and IL-8 in response to high concentration of non-transferrin bound iron in RBC (\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e). Therefore, the observed higher WBC in the old packed RBC transfused patients in this study might indicate the higher level of inflammation in these patients compared to those who received fresh packed RBC.\u003c/p\u003e \u003cp\u003eThe overall findings of this study were in favor of using fresh packed RBC transfusion in patients with thalassemia major in order to achieve higher Hb levels and prolong transfusion intervals. These findings were in line with the current recommendations in Japan and the United Kingdom that prioritize the use fresher packed RBC transfusion (\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e). Although cost effectiveness was not assessed in this study, It can be hypothesized that the use of fresh packed RBC might reduce the health and economic burden of thalassemia major by reducing the number of packed RBC transfusion in long term. In Iran the estimated number of patients with beta thalassemia major is 25000 with median estimated life expectancy of 57 years (\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e, \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e). Considering the 3-5-week transfusion interval and the required two packed RBC bags in each transfusion, delaying transfusion for one week may result in the saving of packed RBC units over time. It was previously shown that 20\u0026ndash;60% of the economic burden of beta thalassemia was related to packed RBC transfusion costs (\u003cspan additionalcitationids=\"CR46\" citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e). In a study conducted in Iran in 2018, the yearly cost of beta thalassemia major was 8321.8 United States Dollar (\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e). High packed RBC transfusion frequency was found to attribute to higher serum ferritin and chelation therapy and thus increased economic burden of disease in a multicenter study in Thailand (\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e). Besides the direct costs of packed RBC transfusion, including cross matching, leukoreduction, antibody evaluation, blood group and Rh matching, indirect costs, including the cost of transportation also attribute to the economic burden of thalassemia major (\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e, \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e). Therefore, considering the effect of using fresh packed RBC can have a long term financial and health benefits for patients with beta thalassemia major through reducing the transfusion frequency.\u003c/p\u003e \u003cp\u003eThis study had some limitations. One of the limitations of this study was the lack of long-term follow up of the patients. It would be more important to document the long-term effects of receiving fresh packed RBC compared to old packed RBC in patients with thalassemia major. Another limitation of this study was lack of evaluating inflammatory markers due to financial limitations; therefore, it is recommended that further studies evaluate inflammatory cytokines in relation to Hb levels and transfusion intervals in patients with thalassemia major.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis study reported that fresh packed RBC transfusion might delay the time of next transfusion due to the higher hemoglobin than baseline four weeks after transfusion possibly due to lower inflammatory properties in fresh packed RBC compared to old packed RBC.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eANOVA Analysis of Variance\u003c/p\u003e \u003cp\u003eHb Hemoglobin\u003c/p\u003e \u003cp\u003eIL-6 Interleukin 6\u003c/p\u003e \u003cp\u003eNNT Number Need to Treat\u003c/p\u003e \u003cp\u003ePLT Platelet\u003c/p\u003e \u003cp\u003eRBC Red Blood Cell\u003c/p\u003e \u003cp\u003eTIBC Total Iron-Binding Capacity\u003c/p\u003e \u003cp\u003eTTI Transfusion Transmitted Infection\u003c/p\u003e \u003cp\u003eWBC White Blood Cell\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe study was conducted in accordance with the Helsinki Declaration and was approved by the Ethics Committee of the Tehran University of Medical Sciences (Ethics code: IR.TUMS.MEDICINE.REC.1403.035). Written informed consent was obtained from all patients or their guardians prior to participating in the study. All personal identifiers were deleted from the collected data to ensure anonymity.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll co-authors read and approved the final version of the manuscript and consented for publication.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets used and analyzed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAuthors declare that they have no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was funded by the Tehran University of Medical Sciences (Grant number: 66917).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026rsquo; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors contributed equally to the preparation of the article.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank all those who helped us prepare this article.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eBenz EJ, Sankaran VG, Thalassemia. 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Asian Hematol Res J. 2025;8(4):314\u0026ndash;23.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePrudent M, Tissot J-D, Lion N. In vitro assays and clinical trials in red blood cell aging: Lost in translation. Transfus Apheres Sci. 2015;52(3):270\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGuidet B. Should fresh blood be recommended for intensive care patients? Critical care. (London England). 2010;14(3):158.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBrunskill SJ, Wilkinson KL, Doree C, Trivella M, Stanworth S. Transfusion of fresher versus older red blood cells for all conditions. Cochrane Database Syst Reviews. 2015(5).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHod EA, Zhang N, Sokol SA, Wojczyk BS, Francis RO, Ansaldi D, et al. Transfusion of red blood cells after prolonged storage produces harmful effects that are mediated by iron and inflammation. Blood. 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Vox Sang. 2016;111(1):43\u0026ndash;54.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHancock V, Cardigan R, Thomas S. Red cell concentrate storage and transport temperature. Transfus Med. 2011;21(5):325\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKlein H, Natanson C, Flegel W. Transfusion of fresh vs. older red blood cells in the context of infection. ISBT Sci Ser. 2015;10(Suppl 1):275\u0026ndash;85.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGarc\u0026iacute;a-Roa M, Del Vicente-Ayuso C, Bobes M, Pedraza AM, Gonz\u0026aacute;lez-Fern\u0026aacute;ndez AC, Mart\u0026iacute;n A. Red blood cell storage time and transfusion: current practice, concerns and future perspectives. Blood Transfus = Trasfusione del sangue. 2017;15(3):222\u0026ndash;31.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMykhailova O, Brandon-Coatham M, Durand K, Olafson C, Xu A, Yi QL, et al. Estimated median density identifies donor age and sex differences in red blood cell biological age. Transfusion. 2024;64:705\u0026ndash;15.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePeters AL, Kunanayagam RK, van Bruggen R, de Korte D, Juffermans NP, Vlaar AP. Transfusion of 35-day stored red blood cells does not result in increase of plasma non-transferrin bound iron in human endotoxemia. Transfusion. 2017;57(1):53\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eObonyo NG, Sela DP, White N, Tunbridge M, Sim B, Rachakonda RH, et al. Effects of transfusing older red blood cells on patient outcomes in critical illness: A retrospective cohort study. Vox Sang. 2025;120(5):481\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePriddee N, Pendry K, Ryan KJTM. Fresh blood for transfusion in adults with beta thalassaemia. 2011;21(6):417\u0026ndash;20.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePriddee N, Pendry K, Ryan K. Fresh blood for transfusion in adults with beta thalassaemia. Transfus Med. 2011;21(6):417\u0026ndash;20.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCappellini M-D, Cohen A, Eleftheriou A, Piga A, Porter J, Taher A. Guidelines for the clinical management of thalassaemia. 2013.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShah FT, Sayani F, Trompeter S, Drasar E, Piga A. Challenges of blood transfusions in β-thalassemia. Blood Rev. 2019;37:100588.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHod EA, Zhang N, Sokol SA, Wojczyk BS, Francis RO, Ansaldi D, et al. Transfusion of red blood cells after prolonged storage produces harmful effects that are mediated by iron and inflammation. 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Economic burden of Thalassemia treatment: An analysis from the Vietnam Social Security perspective. PLoS ONE. 2023;18(11):e0293916.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eUchil A, Muranjan M, Gogtay NJ. Economic burden of beta-thalassaemia major receiving hypertransfusion therapy at a public hospital in Mumbai. Natl Med J India.36.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhen X, Ming J, Zhang R, Zhang S, Xie J, Liu B, et al. Economic burden of adult patients with β-thalassaemia major in mainland China. Orphanet J Rare Dis. 2023;18(1):252.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eEmamgholipour S, Ahmadi B, Rajabi AH, Azarkeivan A, Ebrahimi M, Esmaeilzadeh F. Cost-utility of treatment of the patients with Thalassemia Major in Iran %J. J Iran Blood Transfus. 2018;15(4):257\u0026ndash;64.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRiewpaiboon A, Nuchprayoon I, Torcharus K, Indaratna K, Thavorncharoensap M, Ubol BO. Economic burden of beta-thalassemia/Hb E and beta-thalassemia major in Thai children. BMC Res Notes. 2010;3:29.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-pediatrics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"bped","sideBox":"Learn more about [BMC Pediatrics](http://bmcpediatr.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/bped/default.aspx","title":"BMC Pediatrics","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Hematological factors, stored blood, old blood, fresh blood, children, thalassemia","lastPublishedDoi":"10.21203/rs.3.rs-8928118/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8928118/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eIntroduction\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThalassemia major is a severe hemoglobinopathy that requires blood transfusion; however, long term blood transfusion might increase morbidity and mortality. Hemoglobin (Hb) after transfusion determines the need for next transfusion. There is controversy regarding the effect of fresh packed RBC compared to old packed RBC transfusion on Hb. This study compared hemoglobin changes following transfusion between fresh and old packed RBC in transfusion-related thalassemia (TRT) patients.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMaterials and methods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn this cohort study, 140 TRT patients (age \u0026gt; 5years) referred to Bahrami Children's Hospital in mid-2013 were included. Equal number (n = 70) of age and gender matched TRT patients with exposure to fresh (stored for less than 7 days) and old (stored for more than 7 days) packed RBC transfusion were followed for 14 days. In both groups, demographic information (age, sex) were recorded at enrolment. Physical examination data and blood parameters, including red blood cells (RBC), white blood cells (WBC), platelets (PLT), Hb, ferritin, transferrin, total iron binding capacity and bilirubin at baseline and follow up.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTime-group interaction was significant only for WBC (p \u0026lt; 0.001) and Hb (p = 0.029) indicating that fresh packed RBC transfusion resulted in higher Hb and lower WBC at follow up compared to baseline levels. Among the baseline variables only fresh packed RBC transfusion was found to increase the odds of having Hb of at least 1 mg/dL higher than baseline (OR = 15.465, p \u0026lt; 0.001).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFresh packed RBC transfusion might increase the interval between transfusions due to more Hb elevation rate after transfusion in thalassemia major.\u003c/p\u003e","manuscriptTitle":"Comparative study of hemoglobin changes following transfusion of fresh and old blood in patients with thalassemia referred to Bahrami Hospital","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-03-13 17:42:29","doi":"10.21203/rs.3.rs-8928118/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-03-24T06:42:59+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-23T15:53:01+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-21T22:23:34+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-17T18:27:39+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"173473230607820507462422773827327006656","date":"2026-03-17T16:22:42+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"27279525521009671481208104828233178762","date":"2026-03-16T19:44:46+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"89841059687898386235187197884921544024","date":"2026-03-16T13:15:41+00:00","index":"hide","fulltext":""},{"type":"editorInvited","content":"","date":"2026-03-16T08:55:21+00:00","index":"","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-03-10T13:30:29+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-03-10T02:51:23+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-03-09T14:10:16+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Pediatrics","date":"2026-02-27T14:32:26+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-pediatrics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"bped","sideBox":"Learn more about [BMC Pediatrics](http://bmcpediatr.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/bped/default.aspx","title":"BMC Pediatrics","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"6af3366d-999e-4008-babd-1d68b375d8c7","owner":[],"postedDate":"March 13th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-04-29T01:08:11+00:00","versionOfRecord":[],"versionCreatedAt":"2026-03-13 17:42:29","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8928118","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8928118","identity":"rs-8928118","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}