RBC transfusion and necrotizing enterocolitis in very preterm infants: a multicenter observational study (2024)

Necrotizing enterocolitis (NEC) is a frequently encountered gastrointestinal disorder that significantly contributes to adverse outcomes in premature infants. The etiologies of NEC encompass formula feeding, hypoxia, infection¹. However, the debate over whether red blood cell (RBC) transfusion is associated with NEC in preterm infants continues to be widely discussed².

Despite decades of extensive research, the relationship between blood transfusion and NEC remains elusive. Studies have conflicting findings^{3,4,5,6,7,8,9,10,11}, with some suggesting RBC transfusion as a cause of intestinal injury, others proposing it as protective against NEC, and some finding no link. These evidences supporting this association is of lower quality, primarily consisting of case–control studies, with only a limited number of cohort studies conducted thus far. Additionally, the restricted sample sizes across all these studies pose a challenge to definitively clarify the connection between transfusion and NEC. Does transfusion-associated necrotizing enterocolitis (TANEC) exist, and if so, at what critical time point does it occur? Furthermore, what is the prognosis of infants affected by TANEC? A more comprehensive understanding of association between transfusion and NEC is urgently needed and will greatly benefit disease prevention and prognosis.

This study, based on data from the Chinese Neonatal Network (CHNN) cohort study involving very premature infants, seeks to establish a causal link between transfusions and NEC. Additionally, it aims to investigate the prognosis of TANEC infants. The study will provide a more detailed description of TANEC and serve as a basis for disease prevention and treatment.

Study design

This retrospective study conducts a secondary analysis of the CHNN database. Established in 2018, this collaborative network encompasses 79 (by 2021) neonatal intensive care units (NICUs) across China. The network collects clinical data on infants with a birth weight of < 1500g or a gestational age of < 32weeks from member hospitals¹². The study was undertaken with the approval from the ethics review board of the Children’s Hospital of Fudan University (2018–296), the central hub of CHNN. Written informed consent was not required for retrospective observational study as per the [Ethics Review Board of the Children’s Hospital of Fudan University]. All methods in this study were performed in accordance with the relevant guidelines and regulations.

Study population

This study utilized the CHNN database to identify all infants who received RBC transfusions between January 2019 and December 2021. Transfusion strategy was implemented in accordance with the "Guideline for Pediatric Transfusion" issued by the National Health Commission of the People's Republic of China¹³. Infants with major congenital anomalies or missing data regarding NEC or receipt of RBC transfusions were excluded from the analysis.

Definitions

NEC was defined as Bell’s Stage II or greater according to an established criteria¹⁴. Spontaneous Intestinal Perforation (SIP) refers to individuals diagnosed intraoperatively or imaging suggestive of intestinal perforation lacking clinical features of necrotizing enterocolitis (NEC) is not included in the NEC group. Infants who received transfusions but did not develop NEC during hospitalization were categorized “No-NEC”. Infants with NEC were segregated into post-transfusion NEC, pre-transfusion NEC and Undefined NEC (Ud-NEC) based on the sequence of NEC and transfusion. Ud-NEC referred to cases where transfusion and NEC occurred on the same day and the onset of NEC (before or after transfusion) was unclear. Among the post-transfusion NEC, NEC that occurred within two calendar days after transfusion exposure was defined as TANEC, NEC cases that occurred beyond 2days post transfusion, along with pre-transfusion NEC cases, were considered unrelated to transfusion and defined as UNTA-NEC. The time interval between transfusion and NEC refers to the period between the most recent transfusion and the onset of NEC. The accumulated number of transfusions refers to the total count of transfusions administered before TANEC.

Variables

Trained data abstractors obtained variables from the neonate's medical records¹⁵. The data were then transmitted electronically to central hub of CHNN, maintaining patient anonymity. At each site, site investigators were accountable for data quality control. Routine data auditing, and periodic feedback were provided to each site to ensure data quality ¹⁶.

Clinical characteristics of TANEC infants were detailed including maternal age, cesarean delivery, birth weight, gestational age by best obstetric estimate, sex, small for gestational age(SGA, birth weight < 10th percentile for the gestational age according to the Chinese neonatal birth weight values¹⁷), placenta transfusion including delayed cord clamping and umbilical cord milking, low 5-min Apgar score (Apgar score ≤ 7), endotracheal intubation during resuscitation, inborn, placental transfusion, age at the 1st feed and duration of fasting and antibiotic therapy in first seven days of life. Age at transfusion was defined as the postnatal days of life (DOL) and postmenstrual age (PMA).

The outcomes of TANEC infants included cystic periventricular leukomalacia (PVL), severe retinopathy of prematurity (ROP), late-onset sepsis (LOS), severe bronchopulmonary dysplasia (BPD), and mortality. Cystic PVL was defined as the presence of periventricular cysts on cranial ultrasound or magnetic resonance imaging. Severe ROP was defined as ROP stage ≥ 3 according to the International Classification of ROP¹⁸. LOS was defined as culture-proven sepsis between 3 and 28 calendar days of life. Severe BPD was defined as nasal continuous positive airway pressure or intermittent positive pressure ventilation or invasive ventilation requirement at 36weeks of PMA or at discharge, transfer or death if before 36weeks corrected gestational age¹⁹.

Statistical analysis

Data analyses were conducted using SAS version 9.4 (SAS Institute; Cary, NC, USA). Categorical variables were presented as frequencies and percentages, and group comparisons were performed using the chi-square test. Non-normally distributed data were presented as medians and quartiles, and group comparisons were performed using the Wilcoxon test. The normality of the time interval was assessed using the Shapiro–Wilk test. If W = 1 and P > 0.05, it suggests a normal distribution. Otherwise, a non-normal distribution is indicated. Multiple logistic regression analysis was performed to determine the odds ratio (OR) for TANEC for each clinical factor, with adjustment for confounding variables that showed baseline imbalances among the groups. Statistical significance was defined as a two-tailed P < 0.05.

Ethical approval

Ethics review board of the Children’s Hospital of Fudan University (2018-296) and all participating hospitals.

Characteristics of included cases

Between Jan 2019 and Dec 2021, a total of 31915 very preterm infants with a gestational age < 32weeks or a birth weight < 1500g were identified. Among 16,494 patients who received RBC transfusions during their hospitalization, we recorded a total of 41,973 transfusion episodes and 1281 cases of NEC (7.7% as opposed to 4.9% across all infants) were recorded. Of all NEC cases, there were 535 pre-transfusion NEC, 337 Ud-NEC and 409 post-transfusion NEC (Fig.1).

Distribution of time intervals between transfusion and NEC

The distribution curve of time intervals between transfusion and NEC was plotted. Of the 409 post-transfusion NEC cases identified, 149 (36.4%) occurred within 2days after transfusion, 201 (49.1%) within 3days, 224 (54.8%) within 4days, and 282 (68.9%) within one week. With an increase in the time elapsed after transfusion, there was a reduction in the occurrence of NEC (Fig.2 and Table S1). The time interval between transfusion and the presentation of NEC exhibits a non-normal distribution (Shapiro–Wilk test; W = 0.513, P < 0.001), suggesting an association between transfusion and NEC.

Prognosis of TANEC infants compared with No-NEC or UNTA-NEC infants

Characteristics of infants of TANEC, UNTA-NEC, and No-NEC are shown in Table 1. TANEC primarily occurred between DOL15 to 42 (60.4% of all TANEC, Table S2) or at 31 to 34 weeks of PMA (59.7% of all TANEC, Table S3). By comparing with No-NEC, infants of TANEC exhibited a higher mortality rate (adjusted OR 1.69; 95% CI 1.08 to 2.64; P = 0.022), a higher occurrence of severe BPD (adjusted OR 2.03; 95% CI 1.41 to 2.91; P < 0.001) and more frequent LOS (adjusted OR 2.06; 95% CI 1.37 to 3.09; P < 0.001). When compared with UNTA-NEC, TANEC infants still showed a higher rate of severe BPD (OR 1.76; 95% CI 1.18 to 2.62; P = 0.006) (Fig. 3 and Table S4).

This hospital-based, large-scale, multicenter observational study, featuring the largest sample size to date, unveils that more than one-third of post-transfusion NEC manifest within 2days following transfusion. Furthermore, TANEC infants are often associated with increased mortality, higher rates of severe BPD and LOS.

The association between RBC transfusion and NEC has consistently been a controversial topic. Some studies assert RBC transfusion as a cause of intestinal injury^3,4,5,6, while others have proposed that transfusion may serve as a protective factor against NEC^7,8 or has no link with NEC^9,10, and that anemia may pose as an independent risk factor for TANEC¹¹. Recent bench studies have uncovered some evidence supporting TANEC. For example, researches have demonstrated that changes in mesenteric blood flow velocity, reoxygenation, and reperfusion after transfusion could incite intestinal oxidative stress injury^20,21,22. Furthermore, multiple immune factors such as IL-1β, IL-6, IFN-γ, and ICAM-1 have been shown to increase in circulation following RBC transfusion^23,24. Studies have described a murine model of NEC instigated by transfusions after anemia, illustrating typical NEC-like gut injuries in the anemia-transfusion group within 48h post transfusion due to Toll-like receptor-4-mediated injury and intestinal epithelial barrier dysfunction^25,26. Considering our data in tandem with prior animal studies, it is plausible that RBC transfusion is associated with NEC.

Definitive definition of TANEC varied in previous studies, with presenting variable periods ranging from 48h to 7days^11,27,28. In this study, we observed that 36.9% of post-transfusion NEC cases occurred within the first 2days after transfusion, with a marked decrease in incidence thereafter. This finding provides more evidence to support the definition of TANEC as NEC occurring within 48h following RBC transfusion.

To mitigate the occurrence of TANEC, it is crucial to identify the high-risk population and delineate specific transfusion characteristics associated with TANEC. Our study suggests that very preterm infants at 15–42 DOL or at 31–34weeks may be more susceptible to developing TANEC, and thus could potentially benefit from preventive measures such as considering withholding feeding during transfusions^29,30. It is also possible that the underlying cause is the high incidence of anemia during this period, which requires more blood transfusions.

This large retrospective study provides detailed characteristics of transfusions associated with a higher risk of NEC, however, it is important to acknowledge several limitations. First, the retrospective design of the study precluded the analysis of potential factors such as hemoglobin levels, the volume of transfusion, and feeding volume during transfusion. Fortunately, all member units of CHNN adhere to the "Guideline for Pediatric Transfusion" issued by the National Health Commission of the People's Republic of China, thereby effectively mitigating the impact of these factors to the maximum extent possible. Second, the exclusion of transfusion episodes without a definite time interval with Ud-NEC may have resulted in selection bias, although this was necessary for accurate patient grouping. Nevertheless, the study's findings remain robust.

Author notes

1. A list of authors and their affiliations appears at the end of the paper.

Authors and Affiliations

1. Department of Neonatology, Children’s Medical Center, First Hospital of Jilin University, No. 1 Xinmin Street, Changchun, 130021, Jilin, China

   Dan Dang&Hui Wu

2. Department of Neonatology, Children’s Hospital of Fudan University, 399 Wanyuan Road, Minhang District, Shanghai, 201102, China

   Xinyue Gu,Siyuan Jiang,Wenhao Zhou,Yun Cao&Jianguo Zhou

3. Department of Neonatology, The Third Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, China

   Wenli Li

4. Department of Pediatrics, Maternal-Infant Care Research Centre, Mount Sinai Hospital, University of Toronto, Toronto, ON, Canada

   Shoo Kim Lee&Shoo Kim Lee (Chairmen)

5. Children’s Hospital of Fudan University, Shanghai, China

   Chao Chen

6. Children’s Hospital of Zhejiang University School of Medicine, Zhejiang, China

   Lizhong Du (Vice-Chairmen)

7. The Third Affiliated Hospital of Zhengzhou University, Zhengzhou, China

   Xiuyong Chen

8. Guangzhou Women and Children’s Medical Center, Guangzhou, China

   Huyan Zhang

9. Tianjin Obstetrics & Gynecology Hospital, Tianjin, China

   Xiuying Tian

10. Gansu Provincial Maternity and Child Care Hospital, Lanzhou, China

    Jingyun Shi

11. Northwest Women’s and Children’s Hospital, Xi’an, China

    Zhankui Li

12. Shenzhen Maternity and Child Health Care Hospital, Shenzhen, China

    Chuanzhong Yang

13. Guizhou Women and Children’s Hospital, Guiyang, China

    Ling Liu

14. Suzhou Municipal Hospital Affiliated to Nanjing Medical University, Nanjing, China

    Zuming Yang

15. Shengjing Hospital of China Medical University, Shenyang, China

    Jianhua Fu

16. Children’s Hospital of Shanxi, Taiyuan, China

    Yong Ji

17. Quanzhou Women and Children’s Hospital, Quanzhou, China

    Dongmei Chen

18. Fujian Women and Children’s Medical Center, Guangzhou, China

    Changyi Yang

19. Children’s Hospital of Nanjing Medical University, Nanjing, China

    Rui Chen

20. Hunan Children’s Hospital, Changsha, China

    Xiaoming Peng

21. Qingdao Women and Children’s Hospital, Qingdao, China

    Ruobing Shan

22. Nanjing Maternity and Child Health Care Hospital, Nanjing, China

    Shuping Han

23. The First Bethune Hospital of Jilin University, Changchun, China

    Hui Wu

24. The First Affiliated Hospital of Anhui Medical University, Hefei, China

    Lili Wang

25. Women and Children’s Hospital of Guangxi Zhuang Autonomous Region, Nanning, China

    Qiufen Wei

26. The First Affiliated Hospital of Xinjiang Medical University, Ürümqi, China

    Mingxia Li

27. Foshan Women and Children’s Hospital, Foshan, China

    Yiheng Dai

28. The Affiliated Hospital of Qingdao University, Qingdao, China

    Hong Jiang

29. Henan Children’s Hospital, Zhengzhou, China

    Wenqing Kang

30. Children’s Hospital of Shanghai, Shanghai, China

    Xiaohui Gong

31. Chongqing Health Care Center for Women and Children, Chongqing, China

    Xiaoyun Zhong

32. Children’s Hospital of Chongqing Medical University, Chongqing, China

    Yuan Shi

33. Wuxi Maternity and Child Healthcare Hospital, Wuxi, China

    Shanyu Jiang

34. Children’s Hospital of Soochow University, Suzhou, China

    Bing Sun

35. People’s Hospital of Xinjiang Uygur Autonomous Region, Ürümqi, China

    Long Li

36. Yuying Children’s Hospital Affiliated to Wenzhou Medical University, Wenzhou, China

    Zhenlang Lin

37. Shanghai First Maternity and Infant Hospital, Shanghai, China

    Jiangqin Liu

38. Anhui Provincial Hospital, Hefei, China

    Jiahua Pan

39. Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, Shanghai, China

    Hongping Xia

40. Qilu Children’s Hospital of Shandong University, Dezhou, China

    Xiaoying Li

41. The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China

    Falin Xu

42. General Hospital of Ningxia Medical University, Yinchuan, China

    Yinping Qiu

43. Hebei Children’s Hospital, Shijiazhuang, China

    Li Ma

44. Hainan Women and Children’s Hospital, Haikou, China

    Ling Yang

45. The Second Xiangya Hospital of Central South University, Changsha, China

    Xiaori He

46. Ningbo Women & Children Hospital, Ningbo, China

    Yanhong Li

47. Xiamen Children’s Hospital, Xiamen, China

    Deyi Zhuang

48. Shaanxi Provincial People’s Hospital, Xi’an, China

    Qin Zhang

49. The Affiliated Hospital of Southwest Medical University, Luzhou, China

    Wenbin Dong

50. Shanghai Children’s Medical Center Affiliated to Shanghai Jiaotong University School of Medicine, Shanghai, China

    Jianhua Sun

51. First Affiliated Hospital of Kunming Medical University, Kunming, China

    Kun Liang

52. Changzhou Maternal and Children Health Care Hospital, Changzhou, China

    Huaiyan Wang

53. Shenzhen Children’s Hospital, Shenzhen, China

    Jinxing Feng

54. Jiangxi Provincial Children’s Hospital, Nanchang, China

    Liping Chen

55. Xiamen Maternity and Child Health Care Hospital, Xiamen, China

    Xinzhu Lin

56. Zhuhai Center for Maternal and Child Health Care, Zhuhai, China

    Chunming Jiang

57. Guangdong Women and Children’s Hospital, Guangzhou, China

    Chuan Nie

58. Wuhan Children’s Hospital, Wuhan, China

    Linkong Zeng

59. Beijing Children’s Hospital of Capital Medical University, Beijing, China

    Mingyan Hei

60. Maternal and Children Hospital of Shaoxing, Shaoxing, China

    Hongdan Zhu

61. The First People’s Hospital of Yunnan Province, Kunming, China

    Hongying Mi

62. Dehong People’s Hospital of Yunnan Province, Dehong, China

    Zhaoqing Yin

63. First Affiliated Hospital of Xian Jiaotong University, Xi’an, China

    Hongxia Song

64. Inner Mongolia Maternal and Child Health Care Hospital, Xilinhot, China

    Hongyun Wang

65. Dalian Municipal Women and Children’s Medical Center, Dalian, China

    Dong Li

66. Lianyungang Maternal and Children Health Hospital, Lianyungang, China

    Yan Gao

67. Children’s Hospital Affiliated to Capital Institute of Pediatrics, Beijing, China

    Yajuan Wang

68. Anhui Children’s Hospital, Hefei, China

    Liying Dai

69. Fuzhou Children’s Hospital of Fujian Province, Fuzhou, China

    Liyan Zhang

70. Kunming Children’s Hospital, Kunming, China

    Yangfang Li

71. Shenzhen Hospital of Hongkong University, Shenzhen, China

    Qianshen Zhang

72. Peking Union Medical College Hospital, Beijing, China

    Guofang Ding

73. Obstetrics & Gynecology Hospital of Fudan University, Shanghai, China

    Jimei Wang

74. The Affiliated Hospital of Guizhou Medical University, Guizhou, China

    Xiaoxia Chen

75. Qinghai Women and Children Hospital, Qinghai, China

    Zhen Wang

76. The International Peace Maternity & Child Health Hospital of China Welfare Institute, Shanghai, China

    Zheng Tang

77. Children’s Hospital of Zhejiang University, Zhejiang, China

    Xiaolu Ma

78. Inner Mongolia People’s Hospital, Hohhot, China

    Xiaomei Zhang

79. Xiamen Humanity Hospital, Xiamen, China

    Xiaolan Zhang

80. Shanghai General Hospital, Shanghai, China

    Fang Wu

81. The First People’s Hospital of Yinchuan, Yinchuan, China

    Yanxiang Chen

82. The Third Hospital of Nanchang, Nanchang, China

    Ying Wu

83. University of Alberta, Alberta, Canada

    Joseph Ting

Consortia

Contributions

Dan Dang: performed study concept and design, performed development of methodology, analysis and interpretation of data, writing – original draft, read and approved the final paper. Xinyue Gu: analysis and interpretation of data, statistical analysis, read and approved the final paper. Wenli Li: analysis and interpretation of data, read and approved the final paper Siyuan Jiang: performed study concept and design, provided technical and material support, read and approved the final paper. Wenhao Zhou, Yun Cao and Shoo Kim Lee: Create and maintain CHNN data platform, read and approved the final paper. Hui Wu and Jianguo Zhou: performed study concept and design, project administration, writing – review & editing, read and approved the final paper.

Corresponding authors

Correspondence to Hui Wu or Jianguo Zhou.

Competing interests

The authors declare no competing interests.

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