Efficacy and safety of high and low dose recombinant human erythropoietin on neurodevelopment of premature infants: A meta-analysis.

BACKGROUND: To evaluate the effect of recombinant human erythropoietin (rhEPO) in nervous system of premature infants including different dosage.
METHODS: The multiple databases like Pubmed, Embase, Cochrane databases and China National Knowledge Database were used to search for the relevant studies, and full-text articles involved in the evaluation on effect of rhEPO for neurodevelopment among premature infants. Review Manager 5.2 was adopted to estimate the effects of the results among selected articles. Forest plots, sensitivity analysis and bias analysis for the articles included were also conducted.
RESULTS: Finally, 10 eligible studies were eventually satisfied the included criteria. The results showed that rhEPO was much higher than placebo group in composite cognitive score (MD = 5.89, 95% confidential interval {CI} [1.95, 9.82], P = .003; I2 = 89%), there was no significant difference between rhEPO and placebo groups (RR = 0.93, 95% CI [0.60, 1.43], P = .74; I2 = 51%) and no difference in neurodevelopmental impairment between rhEPO and placebo was insignificant (RR = 0.55 95% CI [0.30, 1.02], P = .06). Composite cognitive score in high dose rhEPO was much higher than placebo group (MD = 10.39, 95% CI [8.84, 11.93], P < .0001, I2 = 0%) and low dose rhEPO also had higher composite cognitive score than placebo group (MD = 2.58, 95% CI [0.80, 4.37], P = .004, I2 = 11%). Limited publication bias was observed in this study.
CONCLUSION: Recombinant human erythropoietin might be a promotor for neurodevelopment among premature infants with limited adverse events.

Introduction

In developed countries, premature babies accounted for 5% to 7% of live births. Due to economic factors, developing countries had a higher premature birth rate than developed countries. The risk of delayed nervous system development in preterm infants was significantly higher than that in term infants. Neurodevelopmental disorders (nervous system abnormalities including cerebral palsy, deafness, blindness and/or mental development index score <70) accounted for 36% to 48% of preterm infant survivors.[ Studies had shown that the nursing progress of babies born before 28 weeks of pregnancy was related to a significant increase in survival. However, about 40% of infants born before 28 weeks would have one or more serious injuries (such as cerebral palsy, intellectual disability, deafness or blindness) and the increase would be higher after a few weeks.[ Children were at higher risk of behavioral autism and mental disorders. There was an urgent need for neuroprotective agents to improve the prognosis of preterm infants.

Erythropoietin (EPO) was originally used to promote red blood cell production and treat anemia.[ Animal experiments showed that erythropoietin and its receptors are expressed in the central nervous system, and that erythropoietin could pass through the blood-cerebrospinal fluid barrier of many animals to play a protective role in the nervous system. Various animal brain injury models such as hypoxic-ischemic brain injury, focal cerebral ischemia, traumatic injury of the brain and spinal cord, autoimmune cerebrospinal meningitis, showed that EPO treatment could reduce brain injury and improve Nerve function.[

Although the prognosis of premature babies had improved in recent decades, they still experienced severe long-term neurodevelopmental delay.[ Magnetic resonance imaging (MRI) showed that the underlying pathology was called premature encephalopathy. Among several drug candidated to prevent brain damage or promote brain development, erythropoietin (EPO) had proven to be the most promising.[ Erythropoietin stimulated the proliferation, survival and differentiation of erythroid cells. Recent evidence indicated that EPO played a role in tissue development and protection, and EPO receptors were also expressed in other cell types such as endothelial cells, glial cells and neuronal cells.[ Animal experiments first proved the neuroprotective effect of EPO. The underlying mechanisms explaining this effect included inhibition of glutamate release, regulation of intracellular calcium metabolism, production of anti-apoptotic factors, and anti-inflammatory and antioxidant effects. Preclinical data supported the results of clinical trials to evaluate the neuroprotective effect of recombinant human erythropoietin (rhEPO) on term infants with hypoxic-ischemic encephalopathy. Retrospective recent studies using rhEPO had shown that EPO also improved the neurodevelopmental outcome of preterm infants.[ When passing through the blood-brain barrier, epo must be administered at a high dose of 2000 to 5000 IU/kg body weight. Premature infants tolerate these high doses well. Compared with reports several weeks after treatment, treatments sufficient to block brain receptors in a short period of time did not increase the risk of retinopathy in premature infants.[ According to reports, there was an association between early high-dose rhEPO and full-term brain MRI evaluation of very premature infants, and there was an association between the incidence of white and gray matter damage.[

In recent years, the value of rhEPO in neurodevelopment among premature infants had been noted, but the detailed role of rhEPO had not been fully understood. Here, we conducted a meta-analysis to evaluate the effects of rhEPO in neurodevelopment among premature infants.

Methods

Literature search strategy

We searched articles published between January 2010 and March 2020 for rhEPO and placebo in premature infants neurodevelopment. Searchable databases included PubMed, EmBase, Cochrane database and China National Knowledge database, and use the following strategy: (neurodevelopment OR nerve∗ OR neurological∗) AND (recombinant human erythropoietin OR rhEPO) AND (dose OR different dose). There were no restrictions on the publication language in the literature search. In order to maximized the specificity and sensitivity of the search, the author should also check the reference list of research to seek other relevant research that was not found through the search strategy. Besides, an ethics committee or institutional review board approved the study.

inclusion criteria and exclusion criteria

Study selection

We used the following inclusion criteria for our research:

A study with RCT design;

A study to evaluate rhEPO and placebo in clinical effects and safety;

Containing data in different year;

Available in full text.

If the research meets one of the following conditions, it is excluded:

Overlapping data or overlapping review articles;

The included samples were not premature infants; and

Other articles design.

Data extraction and quality assessment

Two commentators independently scanned the full text of the manuscript and extracted the following data from each eligible study: first authors name, patient's age and gender, country of origin, year of publication, sample size, study period of each article. Cochrane risk of bias assessment tool was used to evaluate the methodological quality of the study. Cochrane risk assessment tool was a comprehensive tool to consider multiple biases.

Statistical analysis

Review Manager (version 5.2, Cochrane Collaboration, 2011) was used to assess the impact of results in selected reports. For continuous results, the mean difference was calculated by the average difference. Heterogeneity in research I Statistics, a quantitative measure of inconsistency in research. 25% to 50% of the studies with I were considered as low heterogeneity, 50% to 75% of the studies with I were considered as medium heterogeneity, and 75% of the studies with I > 75% were considered as high heterogeneity. If I > 50%, the potential sources of heterogeneity were examined by sensitivity analysis, which omits 1 study in each round and investigates the impact of a single study on portfolio estimation. In addition, when heterogeneity was observed, the random effect model was used, and when it does not exist, the fixed effect model was used. Funnel charts, Begger test and Egger test were used to check for potential publication bias.

Since our research is a meta-analysis, all the data were extracted from published articles, ethical review was unnecessary.

Results

Search process

The electronic search ended with 401 articles. After careful reading, 98 papers had reached the preliminary standard. In the further screening, 88 articles were excluded because of improper research type and insufficient data and article type. Finally, 10 papers were selected for analysis. Figure 1 was a flowchart of identification, inclusion, and exclusion, reflecting the search process and the reason for exclusion.

Figure 1

Flow diagram of the study selection.

Characteristics of included studies

Detailed characteristics of the included studies were presented in Table 1.[ All these studies were published from 2010 to 2020. The sample size ranged from 53 to 1285. Totally 2368 patients were in rhEPO group, and 2265 patients were in placebo group.

Table 1

Characteristics of studies included in the meta-analysis.

Study	Year	Language	Country	cumulative dose of Erythropoietin	Gestational age (weeks)	Groups	n	Years of onset
Fauchere[13]	2015	English	Switzerland	3000 U/kg	29 ± 1.1	rhEPO	229	September 2005 to December 2012
					29 ± 1.2	Placebo	214
Juul[14]	2020	English	America	6000 U/kg	29.1 ± 6.2	rhEPO	476	December 2013 to September 2016
					28.8 ± 6.2	Placebo	460
Leuchter[15]	2014	English	Switzerland	3000 U/kg	29.5 ± 1.5	rhEPO	256	2005 to 2012
					29 ± 1.6	Placebo	239
Luciano[16]	2015	English	Italy	6300 U/kg	28.1 ± 1.7	rhEPO	59	December 2004 to January 2007
					28.3 ± 1.2	Placebo	45
Natalucci[17]	2016	English	Switzerland	3000 U/kg	29.2 ± 1.6	rhEPO	191	2005 to 2012
					29.3 ± 1.6	Placebo	174
Neubauer[18]	2010	English	Germany	8574 U/kg	27.1 ± 1.9	rhEPO	89	January 1993 to December 1998
					27.2 ± 2.1	Placebo	57
Ohls[19]	2014	English	America	1200 U/kg	27.8 ± 1.9	rhEPO	29	2008 to 2012
					27.8 ± 1.6	Placebo	24
Qiao[20]	2017	English	China	1600 U/kg	31.3 ± 1.5	rhEPO	32	February 2014 to June 2014
					30.3 ± 2.1	Placebo	31
Song[21]	2016	English	China	3500 U/kg	30.4 ± 1.4	rhEPO	366	January 2009 to June 2013
					30.4 ± 1.3	Placebo	377
Wang[22]	2020	English	China	3500 U/kg	29.7 ± 1.5	rhEPO	641	January 2014 to June 2017
					30 ± 1.6	Placebo	644

Assessment of the quality of the included studies: low risk of bias (green hexagons), unclear risk of bias (yellow hexagons), and high risk of bias (red hexagons).

Quality assessment of included studies.

Results of quality assessment

The Cochrane risk of bias assessment tool was used to evaluate the risk of patient selection problems in 10 trials. None showed the problem of bias. In view of the bias summary, there was no problem in selection bias, performance bias, detection bias, attrition bias and reporting bias. In general, 10 included trials had no risk of bias.

Results of meta-analysis

Meta-analysis about composite cognitive score among premature infants

As shown in Figure 4, 7 included studies involved in composite cognitive score, and there was significant difference between rhEPO and placebo in composite cognitive score. Since I = 89%, random effects model was adopted. The result showed that rhEPO was much higher than placebo group in composite cognitive score (MD = 5.89, 95% confidential interval {CI} [1.95, 9.82], P = .003; I = 89%, fig 4).

Figure 4

Forest plots of composite cognitive score between rhEPO and placebo among premature infants.

Meta-analysis about death among premature infants

Three included studies were involved in death between rhEPO and placebo. As shown in the forest plot (Fig. 5), and since I = 51%, random effects model was adopted. The result of meta-analysis showed that there was no significant difference between rhEPO and placebo groups (RR = 0.93, 95% CI [0.60, 1.43], P = .74; I = 51%).

Figure 5

Forest plots of death between rhEPO and placebo among premature infants.

Meta-analysis about neurodevelopmental impairment among premature infants

In the analysis, 2 articles were included. The results of heterogeneity test showed that random effect model wad was needed to analyze the data (I = 74%). The overall difference of neurodevelopmental impairment between rhEPO and placebo was insignificant (Fig. 6, RR = 0.55 95% CI [0.30, 1.02], P = .06).

Figure 6

Forest plots of neurodevelopmental impairment between rhEPO and placebo among premature infants.

Meta-analysis about subgroup in composite cognitive score

As shown in Figure 7, 3 included studies were involved in high dose rhEPO, and 4 articles were selected in low dose rhEPO. Since I = 0%, fixed effects model was adopted. The result showed that composite cognitive score in high dose rhEPO was much higher than placebo group (MD = 10.39, 95% CI [8.84, 11.93], P < .0001, I = 0%, fig. 7) and low dose rhEPO also had higher composite cognitive score than placebo group (MD = 2.58, 95% CI [0.80, 4.37], P = .004, I = 11%, fig. 7).

Figure 7

Forest plots of subgroup in composite cognitive score between rhEPO and placebo among premature infants.

Results of sensitivity analysis and publication bias

According to meta-analysis, the heterogeneity of composite cognitive score was low (I = 89%). As shown in Figure 8 the heterogeneity of composite cognitive score might be attributed to the different results of each study. When the article of July in 2020 was excluded, I changed to 41%. This indicated that the result in this article was robust.

Figure 8

Sensitivity analysis of composite cognitive score between rhEPO and placebo among premature infants.

A funnel plot for composite cognitive score was performed. Seven studies were included in the plot. The result indicated that there existed limited publication bias since the symmetrical characteristic of the funnel plot is fine, which showed there exist no publication bias in this analysis (Fig. 9). The result of Begger test also suggested that no significant evidence of potential publication bias existed (z = 1.25, P = .131) and Egger test also suggested that no significant evidence of potential publication bias existed (t = 1.21, P = .225).

Figure 9

Funnel plot of publication bias.

Discussion

The results showed that rhEPO was much higher than placebo group in composite cognitive score, which indicated that rhEPO might have the clinical benefits of neurodevelopment. Rao stated that early application of rhEPO can protect the nervous system of preterm infants, and the effect of low-dose application is more effective.[ However, a number of multicenter randomized controlled clinical studies are needed to further confirm the effects of rhEPO. Our results showed that the differences in death and neurodevelopment between rhEPO and placebo had no significance. In the further analysis, we divided samples into 2 subgroups based on dosage of rhEPO. Compared with Zhang's report[ that the effect of low-dose application was more effective than high-dose, the results showed that both rhEPO in high dose and low dose had higher composite cognitive score than placebo group. This was may be caused by different included indicators, and in his article, neonatal behavior neuro-developmental assessment (NBNA), mental development index (MDI), and psychomotor development index (PDI) were analyzed.

Brain damage in premature infants could cover various diseases such as hypoxic-ischemic brain injury, cerebral hemorrhage, hydrocephalus, periventricular white matter softening, etc. Children manifested with cerebral palsy and neurodevelopmental disorders, which could affect movement, behavior, and cognition. In this respect, it seriously affected the quality of life of the children and brings a heavy burden to the family and society.[ The current treatment methods for this disease mainly include mild hypothermia therapy, scavenging free radicals to protect damaged cells, and hormones but the effects are not satisfactory.[ Studies have pointed out that premature stimulation and overtraining of premature infants can have an adverse effect on the central nervous system.[ Therefore, for premature babies, especially those with brain damage, it was difficult to conduct interventional training before correcting the gestational age of 40 weeks. In recent years, early intervention for premature infants with brain injury to improve the quality of life and reduce the occurrence of sequelae had become the focus of neonatal medical research.

Premature babies were often immature, and the immaturity of the nervous system and its complications had an important influence on the long-term development of premature babies. Numerous experiments and retrospective clinical studies had shown that EPO had a similar effect as a neuroprotective factor, and its mechanism might deal with these 2 processes at the same time.[ EPO inhibited the release of glutamate, regulates intracellular calcium metabolism, and induces neuronal apoptosis. Mechanisms might include death factors, reduction of inflammation, reduction of NO-mediated damage, and direct anti-oxidation to prevent acute injury. On the other hand, EPO affected its developmental mechanism by promoting the proliferation and differentiation of preoligodendrocytes, then it stimulated growth factors and Inhibit nerve cell apoptosis.[

EPO was essentially a glycoprotein, which was secreted and released by the kidneys, but it was also expressed in small amounts in brain and other tissues. It was named for its effective function of promoting bone marrow hematopoiesis. It was used in the treatment of renal anemia in the early stage. With the deepening of research, researchers found that it had a certain repair effect on the damage of the nervous system, and could protect the nerve by inhibiting the degradation of nerve cell protein and activating the function of mitochondria.[

In conclusion, recombinant human erythropoietin might be a promotor for neurodevelopment among premature infants with limited adverse events. Besides, some limitations existed in this article. Firstly, the comparison in different areas was not considered, which could be evaluated in the further research. Secondly, the details about complications were not included, and details could be evaluated in the future.

Author contributions

Conceptualization: Na Qin.

Data curation: Huibin Qin, Na Qin.

Formal analysis: Huibin Qin.

Methodology: Na Qin.

Resources: Na Qin.