Role of Citicoline in Neonates with Hypoxic-Ischaemic Encephalopathy

By Saqib Nawaz¹, Arshad Khushdil¹, Sajid Ali Shah¹, Bilal Ahmed Malik¹, Fiza Wali²

Affiliations

1. Department of Paediatrics, National University of Medical Sciences, Rawalpindi, Pakistan
2. Department of Paediatrics, Riphah International University, Rawalpindi, Pakistan

doi: 10.29271/jcpsp.2026.08.1034

ABSTRACT
Objective: To evaluate the short-term neurological outcomes in neonates with moderate-to-severe hypoxic-ischaemic encephalopathy (HIE) treated with citicoline.
Study Design: A randomised controlled trial.
Place and Duration of the Study: Department of Paediatrics, Pak Emirates Military Hospital, Rawalpindi, Pakistan, from 2024 to 2025.
Methodology: Seventy neonates presenting with moderate or severe HIE, according to the Sarnat scoring system, were enrolled. Participants were allocated into two equal groups (n = 35 each) by using a simple randomisation method. The citicoline-treated group received citicoline in addition to routine care, while the control group received only routine care. Clinical outcomes were compared. Data were analysed using the chi-square test for qualitative variables and the Student’s t-test for quantitative variables. A p-value of <0.05 was considered statistically significant.
Results: Out of 70 neonates, 37 (52.8%) were female, while 33 (47.2%) were male. In the citicoline-treated group, the mean duration of NICU stay was 6.23 ± 3.42 days, compared with 8.46 ± 3.26 days in the control group (p = 0.007). Furthermore, the suck and Moro reflexes were established significantly earlier than the control group, with p-values of 0.001 and 0.002, respectively.
Conclusion: This study demonstrated the neuroprotective effects of citicoline in the treatment of HIE in neonates, with statistically significant earlier establishment of suck and Moro reflexes in the citicoline-treated group compared with the control group.

Key Words: Citicoline, Hypoxic-ischaemic encephalopathy, Neonates, Seizures, Suck reflex.

INTRODUCTION

Hypoxic-ischaemic encephalopathy (HIE) is a leading cause of morbidity and mortality in newborn children, occurring as a result of hypoxia, causing acute or sub-acute brain injury.¹ It is more prevalent in developing countries, with reporting rates of 2.3 to 26.5 per 10,000 live births as compared to developed countries, where reporting rates are 1.5 per 1,000 live births.² Moderate HIE causes 10% mortality in neonates, and 30% are at risk of developing disabilities; whereas, mortality caused by severe HIE is 60%, and almost all neonates suffering from severe  HIE  develop  some  form  of  disability.³

Compared to adults, neonates have a very narrow range of blood pressure (BP), which is responsible for regulating cerebral blood flow. Systemic BP regulates cerebral perfusion after the failure  of  initial  compensatory  mechanisms.
 

Systemic hypoxaemia leading to reduced cerebral blood flow is the primary cause of HIE.⁴ Cerebral oxygen demand is reduced due to the neurotransmitter GABA, which is released in the early phase of brain injury. However, in the later stages of ischaemia, uptake of the excitatory neurotransmitter glutamate is impaired, leading to overactivation of excitatory amino acid receptors. This leads to increased intracellular concentrations of sodium and calcium ions, promoting the synthesis of reactive oxygen species (ROS) and nitric oxide (NO), thereby enhancing excitotoxicity and contributing to the cumulative progression of brain injury. The extent of brain damage depends on the severity and duration of the initial injury, production  of  ROS,  and  rate  of  ongoing  apoptosis.^5,6

Citicoline (cytidine 5-diphosphocholine) serves as a precur-sor for phosphatidylcholine synthesis, which is essential for repairing and stabilising neuronal and mitochondrial memb-ranes damaged during hypoxia-ischaemia.^7-9 Citicoline plays a crucial role in inhibiting different steps of the ischaemic cas- cade.¹⁰ It increases dopamine and acetylcholine neurotransmitters, regenerates injured cell membranes, and increases brain plasticity and repair.^11,12

While therapeutic cooling is the standard of care for HIE, its effectiveness is strictly limited to a narrow window of six hours post-birth and often unavailable in resource-limited settings or tertiary care centres in developing countries.

Given the high burden of HIE in Pakistan, there is a critical need for an accessible pharmacological intervention, such as citicoline, that can provide neuroprotection even when the window for therapeutic cooling has passed. This study aimed to evaluate the short-term neurological benefits of citicoline as an adjunct to routine care in these neonates.

METHODOLOGY

The study is a single-blinded, randomised controlled trial. While the medical team was aware of the intervention for safety monitoring purposes, the parents of the neonates remained blinded to the treatment allocation to minimise bias in reported observations. The study was registered with ClinicalTrials.gov (NCT06522581) and conducted in the neonatal intensive care unit (NICU) of Pak Emirates Military Hospital, Rawalpindi, Pakistan, from 2024 to 2025. A non-probability convenience sampling technique was used. A sample size of 70 neonates was calculated by using the World Health Organisation (WHO) sample size calculator. Using an incidence of 4.5%¹³ as the reference, a 95% confidence interval, and a 5% margin of error, the calculated sample size was 67; therefore, a rounded sample size of 70 was selected. The study included all newborns who presented with moderate or severe HIE according to the Sarnat scoring system and who either did not meet the criteria for therapeutic cooling (postnatal age >6 hours, birth weight <2000 g, and gestational age <36 weeks) or were managed in settings where therapeutic cooling was unavailable. Patients whose parents did not consent to inclusion in the study or who had undergone therapeutic cooling were excluded from this study.

Informed consent was obtained from parents or legal guardians. Patient confidentiality and privacy were strictly maintained. The study adhered to ethical guidelines and was conducted after obtaining approval from the institutional review board. Data were collected using a pre-formatted questionnaire. The questionnaire consisted of the following parts: demographic and clinical data.

A total of 70 patients were categorised into two groups. The citicoline-treated group comprised 35 neonates who received citicoline in addition to supportive measures, including respiratory support, thermoregulation, nasogastric feeding, careful fluid management, avoidance of hypoglycaemia, and treatment of seizures. All neonates in this group received citicoline 15 mg/kg x12 hourly IV until development of the suck reflex. All neonates in the citicoline-treated group tolerated the drug without any adverse effects. The control group had 35 neonates who were managed with the same supportive measures. Patients were categorised according to the Sarnat scoring system used for HIE grading. Factors such as total duration of NICU stay, total number of days on mechanical ventilation or any other type of respiratory support, and need for inotropes were assessed. While the primary outcome of this study was the final status of the neonate (death or discharge), the secondary outcomes included the total duration of stay in the NICU, the total number of days on mechanical ventilation or any other type of respiratory support, the establishment of the Moro and suck reflexes, and the need for inotropes. These patients were assessed for the development of any systemic complications such as respiratory distress syndrome, pulmonary hypertension, gastrointestinal ulceration, necrosis, or perforation, and shock.           

The collected data were analysed using SPSS version 26 for biostatistical analysis. The chi-square test was used for qualitative variables, and the t-test was used for quantitative variables. A p-value <0.05 was considered statistically signifi- cant. A confidence interval of 95% and a margin of error 5% was used. Qualitative variables, including gender, mode of delivery, citicoline administration, development of seizures, respiratory distress syndrome, pulmonary hypertension, gastrointestinal ulceration, necrosis or perforation, and shock, were expressed as frequencies and percentages. Quantitative data such as age, birth weight, APGAR score, total duration of NICU stay, and days on mechanical ventilation were calculated as mean ± SD.

RESULTS

Seventy neonates were included in the study and subdivided into two equal groups: citicoline-treated group (n = 35) and the control group (n = 35). Three neonates, all diagnosed with severe HIE, died during the study; two were from the control group, and one was from the citicoline-treated group. 

One mortality (2.9%) occurred in the citicoline-treated group due to severe neonatal sepsis and severe HIE. The mean gestational age of 35 neonates in the citicoline-treated group was 34.71 ± 2.23 weeks. The mean birth weight was 2597 ± 473 grams. Among the remaining 34 neonates in the citicoline group, six (17.1%) had severe HIE, while 29 (82.9%) had moderate HIE. The mean APGAR scores at one and five minutes were 6.2 ± 0.7 and 7.2 ± 0.8, respectively. The mean first-hour capillary blood pH was 7.23 ± 0.7, and the mean lactate level was 6.76 ± 2.81 mmol/L. Five (14.2%) neonates required mechanical ventilation, and 12 (34.2%) required non-invasive ventilation. The mean duration of  NICU stay was 6.2 ± 3.4 days. Among 35 babies, 6 (17.1%) developed various complications, including 3 (8.5%) who developed pulmonary hypertension and 3 (8.5%) who experienced respiratory distress syndrome. Out of the total 35 babies included in the study, 34 (97.1%) babies in the citicoline-treated group survived despite complications, while 1 (2.9%) baby passed away.

In the control group, the mean gestational age was 34.28 ± 2.66 weeks, and the mean birth weight was 2,608 ± 435 g. Five (14.3%) neonates had severe HIE, and 30 (85.7%) had moderate HIE. The mean APGAR scores at one and five minutes were 6.4 ± 0.7 and 7.3 ± 0.7, respectively. The mean first-hour capillary blood pH was 7.22 ± 0.83, and the mean lactate level was 7.38 ± 2.68 mmol/L. Four (11.4%) neonates required mechanical ventilation, while 10 (28.6%) required non-invasive ventilation. In the control group, 33 (94.3%) neonates survived, and two (5.7%) died, one neonate due to severe HIE and RDS and the second neonate due to severe HIE and sepsis. 

Comparative outcomes regarding the establishment of the suck and Moro reflexes were statistically significant in favour of the citicoline-treated group, with p-values of 0.001 and 0.002, respectively. Detailed results are summarised in Tables I-IV.

Table I: Comparison of demographic and clinical variables between the citicoline treatment and control groups.
 

Variables	Division	Citicoline-treated group n = 35 (100%)	Control group n = 35 (100%)	p-values*
Gender	Male	18 (51.4)	15 (42.8)	0.473
	Female	17(48.5)	20 (57.1)
Mode of delivery	LSCS	8 (22.8)	9 (25.7)	0.780
	SVD	27(77.1)	26 (74.2)
Metabolic derangements	Hypoglycaemia	6 (17.1)	7 (20)	0.578
	Hypocalcaemia	3(8.6)	1 (2.8)
	None	26(74.2)	27 (77.1)
Grade of HIE	Moderate	29 (82.8)	30 (85.7)	0.743
	Severe	6 (17.1)	5 (14.3)
CTG abnormality	Yes	14 (40)	9 (25.7)	0.203
	No	21 (60)	26 (74.2)
*The chi-square test was used to determine the p-value.

Table II: Comparison of neurological recovery and complications.

Variables	Division	Citicoline-treated group 35 (100%)	Control group 35 (100%)	p-values*
Suck reflex	Absent	6 (17.1)	5 (14.3)	0.743
	Weak	29 (82.8)	30 (85.7)
Moro reflex	Absent	6 (17.1)	5 (14.3)	0.743
	Weak	29 (82.8)	30 (85.7)
Suck reflex follow-up	No	1 (2.8)	1 (2.8)	0.001
	Within a day	15 (42.8)	1 (2.8)
	within 3 days	13 (37.1)	15 (42.8)
	within 7 days	4 (11.4)	13 (37.1)
	within 2 weeks	2 (5.7)	5 (14.3)
Moro reflex follow-up	No	1 (2.8)	1 (2.8)	0.002
	within a day	15 (42.8)	1 (2.8)
	within 3 days	13 (37.1)	17 (48.5)
	within 7 days	4 (11.4)	11 (31.4)
	within 2 weeks	2 (5.7)	5 (14.3)
Neurological signs	Hypotonia	30 (85.7)	33 (94.3)	0.232
	Absent reflexes	5 (14.3)	2 (5.7)
*The chi-square test was used to determine the p-value.

Table III: Comparison of clinical interventions and final neonatal outcomes.

Variables	Category	Citicoline-treated group  35 (100%)	Control group 35 (100%)	p-values*
Seizures	Present	9 (25.7)	8 (22.8)	0.780
	Absent	26 (74.2)	27 (77.1)
Mechanical ventilation	Yes	5 (14.3)	4 (11.4)	0.721
	No	30 (85.7)	31 (88.6)
Inotropes given	yes	6 (17.1)	9 (25.7)	0.382
	no	29 (82.8)	26 (74.3)
Fina status	Died	1 (2.8)	2 (5.7)	0.555
	Survived	34 (97.1)	33 (94.3)
Duration of mechanical ventilation	Mechanical Ventilation not required	30 (85.7)	31 (88.6)	0.923
	within 7 days	4 (11.4)	3 (8.5)
	within 2 weeks	1 (2.8)	1 (2.8)
Seizures controlled	Seizures not present (NA)	26 (74.2)	27 (77.1)	0.391
	Not controlled	1 (2.8)	2 (5.6)
	within 3 days	2 (5.7)	0
	within 7 days	5 (14.2)	6 (17.2)
	within 2 weeks	1 (2.8)	0
*The T-test was used to determine the p-value.

Table IV: Comparison of hospitalisation and treatment durations.

Variables	Groups	n	Mean ± SD (days)	p-values
Duration of seizures	Citicoline-treated group	35	0.97 ± 2.0 0.0 ± 0.0	0.006
	Control group	35
Days on non-invasive ventilation	Citicoline-treated group	35	0.94 ± 1.5 1.42 ± 2.3	0.319
	Control group	35
Days on mechanical ventilation	Citicoline-treated group	35	0.91 ± 2.3 0.71 ± 2.0	0.706
	Control group	35
Stay in the NICU (days)	Citicoline-treated group	35	6.22 ± 3.4 8.45 ± 3.2	0.007
	Control group	35
Duration of inotropes	Citicoline-treated group	35	1.37 ± 3.1 1.40 ± 2.7	0.968
	Control group	35

DISCUSSION

HIE remains a significant contributor to neonatal mortality and is a leading cause of long-term neurodevelopmental disabilities in survivors. Therapeutic hypothermia is the only effective neuroprotective modality proven so far for the treat-ment of hypoxic ischaemic encephalopathy.^14,15 However, therapeutic hypothermia requires specific eligibility criteria, including a gestational age of more than 36 weeks, a birth weight of more than 2000 g, and initiation within 6 hours of birth, which limits its applicability. In the present study, because the neonates did not meet the criteria for therapeutic hypothermia, citicoline was administered to the citicoline-treated group. The suck and Moro reflexes were established significantly earlier in this group than in the control group, which was statistically significant. However, no statistically significant differences were observed between the groups regarding the duration of mechanical or non-invasive ventilation, mortality, or the requirement for inotropes.

In a similar study by Salamah et al., involving 80 neonates with moderate-to-severe HIE, a statistically significant reduction in the NICU stay was also observed in the treatment group.¹⁶ Their research featured a more comprehensive scope, employing objective assessments through cranial ultrasound and magnetic resonance imaging (MRI) of the brain. Furthermore, they followed the population for one year and reported superior neurodevelopmental outcomes in the citicoline-treated group.¹⁶

In this randomised controlled trial, significant differences were observed between neonates with moderate-to-severe HIE who received citicoline and those who did not. Neonates in the citicoline-treated group have achieved normal suck and Moro reflexes earlier as compared to other groups, along with faster improvement in other neurological signs, including grasp reflex and plantar reflex.

A pilot study by Khushdil et al. also explored the role of citicoline in neonates with moderate-to-severe HIE.¹⁷ Methodologically, their study included 20 patients, a smaller cohort than the current study (n = 70). Their population had a higher prevalence of severe HIE (65% vs. 17% of this study), which likely reflects differences in inclusion criteria or study setting.

Regarding outcomes, the findings of the present study corro-borate those of Khushdil et al., as both studies demonstrated significantly earlier establishment of the suck reflex in the citicoline-treated group. Both studies noted an almost equal incidence of seizures between the two groups. A key difference in baseline parameters was the mean gestational age (38.25 ± 2.04 weeks in their study vs. 34.71 ± 2.23 weeks in the present study), suggesting that the cohort included more premature infants.

The efficacy of citicoline in controlling seizures and providing neuroprotection has been documented in multiple studies.^17-19 The current findings regarding reduced NICU stay and improved neurological recovery are consistent with previous international trials. Furthermore, while the present study focused on short-term outcomes, such as the establishment of neonatal reflexes, including Moro and suck reflexes, other researchers reported improved long-term neurodevelopmental scores at 12 months, suggesting that early clinical benefits may have lasting implications.^18,19 These results are further supported by another study observing citicoline effects in HIE neonates.²⁰

The limitations of the present study include a relatively small sample size and a lack of long-term follow-up to assess developmental milestones or post-discharge complications. Additionally, follow-up brain imaging was not performed. Consequently, large-scale studies with expanded sample sizes, continuous long-term follow-up, and serial neuroimaging are required to further ascertain the role of citicoline in the treatment of HIE.

CONCLUSION

Citicoline may have a potential adjunctive role in the management of moderate-to-severe HIE, particularly in settings where therapeutic hypothermia is not feasible. Neonates who received citicoline for the treatment of HIE showed statistically significant earlier establishment of the suck and Moro reflexes. All neonates tolerated citicoline without any observed side effects.

ETHICAL APPROVAL:
Ethical approval was obtained from the Ethical Review Committee of Pak Emirates Military Hospital, Rawalpindi, Pakistan (Ref. No. A/28/ERC/05/2024; dated: 2 January 2024).

PATIENTS’ CONSENT:
Informed consent was obtained from all participants.

COMPETING INTEREST:
The authors declared no conflict of interest.

AUTHORS’ CONTRIBUTIONS:
SN, AK, SAS, BAM, FW: Contribution to the study design, acquisition of data, manuscript drafting, and review of the manuscript critically.
All authors approved the final version of the manuscript to be published.

REFERENCES

1. Ranjan AK, Gulati A. Advances in therapies to treat neonatal hypoxic-ischaemic encephalopathy. J Clin Med 2023; 12(20): 6653. doi: 10.3390/jcm12206653.
2. Oliveira RF, Wagner LH, Silva ASD, Rodrigues MCC, Lacerda GCB. Neurological outcomes in neonates treated with therapeutic hypothermia: Challenges in a developing country. Arq Neuropsiquiatr 2024; 82(9):1-8. doi: 10.1055/s-0044-1790575.
3. Shah M, Sakr M, Balasundaram P. Neonatal Therapeutic Hypothermia. [Updated 2025 Sep 14]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2026 Jan-. Available from: httpss://www.ncbi.nlm.nih.gov/books/NBK5 67714/.
4. Tierradentro-García LO, Saade-Lemus S, Freeman C, Kirschen M, Huang H, Vossough A, et al. Cerebral blood flow of the neonatal brain after hypoxic-ischemic injury. Am J Perinatol 2023; 40(5):475-88. doi: 10.1055/s-0041-1731278.
5. Greco P, Nencini G, Piva I, Scioscia M, Volta CA, Spadaro S, et al. Pathophysiology of hypoxic–ischaemic encephalopathy: A review of the past and a view on the future. Acta Neurologica Belgica 2020; 120(2):277-88. doi: 10.1007/ s13760-020-01308-3.
6. Yang M, Wang K, Liu B, Shen Y, Liu G. Hypoxic-ischaemic encephalopathy: Pathogenesis and promising therapies. Mol Neurobiol 2025; 62(2):2105-22. doi: 10.1007/s12035-024- 04398-9.
7. Adibhatla RM, Hatcher JF. Cytidine 5′-diphosphocholine (CDP-choline) in stroke and other CNS disorders. Neurochem Res 2005; 30(1):15-23. doi: 10.1007/s11064-004-9681-8.
8. Cavalu S, Saber S, Ramadan A, Elmorsy EA, Hamad RS, Abdel‐Reheim MA, et al. Unveiling citicoline's mechanisms and clinical relevance in the treatment of neuroinflammatory disorders. FASEB J 2024; 38(17):e70030. doi: 10. 1096/fj.202400823R.
9. Jasielski P, Piedel F, Piwek M, Rocka A, Petit V, Rejdak K. Application of citicoline in neurological disorders: A systematic review. Nutrients 2020; 12(10):3113. doi: 10.3390/nu 12103113.
10. Agarwal A, Vishnu VY, Sharma J, Bhatia R, Garg A, Dwivedi S, et al. Citicoline in acute ischaemic stroke: A randomised controlled trial. PLoS One 2022; 17(5):e0269224. doi: 10. 1371/journal.pone.0269224.
11. Jeong H, Yoon S, Sung YH, Kim J, Lyoo IK, Yurgelun-Todd DA, et al. Effects of cytidine-5′-diphosphate choline on grey matter volumes in methamphetamine-dependent patients: A randomised, double-blind, placebo-controlled study. J Psychiatr Res 2021; 143:215-21. doi: 10.1016/j.jpsychires. 2021.09.006.
12. Gimenez R, Soler S, Aguilar J. Cytidine diphosphate choline administration activates brain cytidine triphosphate: Phosphocholine cytidylytransferase in aged rats. Neurosci Lett 1999; 273(3):163-6. doi: 10.1016/S0304-3940(99)00660-6.
13. Ullah K, Nadir M, Saddozai S, Khan MQ, Ahmad K, Khan Q. Prevalence and determinants of hypoxic-ischaemic encephalopathy in perinatal asphyxia population of district Mardan, Pakistan. Gomal J Med Sci 2022; 20(2):82-8. doi: 10.46903/gjms/20.02.863.
14. Aker K, Stoen R, Eikenes L, Martinez-Biarge M, Nakken I, Haberg AK, et al. Therapeutic hypothermia for neonatal hypoxic-ischaemic encephalopathy in India (THIN study): A randomised controlled trial. Arch Dis Child Fetal Neonatal Ed 2020; 105(4):405-11. doi: 10.1136/archdischild-2019- 317311.
15. Kumar C, Adhisivam B, Bobby Z, Bhat BV. Magnesium sulfate as an adjunct to therapeutic hypothermia in the management of term infants with hypoxic–ischemic encephalopathy: A randomised, parallel-group, controlled trial. Indian J Paediatr 2023; 90(9):886-92.
16. Salamah A, El Amrousy D, Elsheikh M, Mehrez M. Citicoline in hypoxic ischaemic encephalopathy in neonates: A randomised controlled trial. Ital J Pediatr 2023; 49(1):55. doi: 10.1186/s13052-023-01452-5.
17. Khushdil A, Ahmed Z, Ehsan A. Role of citicoline in treatment of moderate to severe birth asphyxia: A pilot project. J Coll Physicians Surg Pak 2021; 31(12):1511-2. doi: 10.29271/ jcpsp.2021.12.1511.
18. Karpova MN, Zin’kovskii KA,  Kuznetsova LV, Klishina NV. Increase of the seizure threshold in C57BL/6 mice after citicoline administration. Bull Exp Biol Med 2015; 158(3):315-7. doi: 10.1007/s10517-015-2750-y.
19. Karpova MN, Kuznetsova LV, Zin’kovskii KA, Klishina NV. Anticonvulsant effects of combined treatment with citicoline and valproate on acute generalised convulsions in rats. Bull Exp Biol Med 2016; 160(4):429-31. doi: 10.1007/s10517-016- 3188-6.
20. Kuznetsova LV, Karpova MN, Zinkovski KA, Klishina NY. Anticonvulsant effects of citicoline and diazepam in pentylenetetrazole-induced convulsions in rats. Patol Fiziol Eksp Ter 2016; 60(4):20-3.