Coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel virus first identified in Wuhan, China, in late 2019 [1]. Since its emergence, COVID-19 has become one of the most imp order Nidovirales, and is an enveloped, positive-sense, single-stranded ribonucleic acid virus characterized by club-shaped spike projections that give the virus its crown-like appearance under electron microscopy [3,4]. Coronaviruses infect both humans and animals and may produce illnesses ranging from mild upper respiratory tract infections to severe and sometimes fatal diseases such as severe acute respiratory syndrome and Middle East respiratory syndrome [5]. SARS-CoV-2 is believed to have originated from an animal source before adapting to efficient human-to-human transmission, and its rapid dissemination led the World Health Organization to officially name the disease COVID-19 in February 2020 [6]. Transmission occurs mainly through respiratory droplets and close contact, although contaminated surfaces and aerosols may also contribute, especially in crowded or poorly ventilated settings [7]. Viral particles are released when infected individuals cough, sneeze, speak, or breathe, and infection occurs when these particles reach the mucous membranes of susceptible persons [8]. The incubation period is usually between 2 and 14 days, with an average of about 4–6 days, and transmission may occur even before the onset of symptoms in some cases [9]. Clinically, COVID-19 presents with variable manifestations, ranging from asymptomatic infection to critical disease, with common symptoms including fever, dry cough, fatigue, myalgia, headache, dyspnea, sore throat, anosmia, nasal obstruction, rhinorrhea, and gastrointestinal disturbances such as nausea, vomiting, and diarrhea [10,11]. Interleukin-6 is produced by a wide range of immune and stromal cells, including macrophages, monocytes, T lymphocytes, B lymphocytes, endothelial cells, and fibroblasts, and its production is stimulated by several pro-inflammatory signals, particularly interleukin-1β and tumor necrosis factor-α [11]. Elevated serum interleukin-6 levels in COVID-19 indicate hyperinflammation and are considered one of the major laboratory abnormalities in critically ill patients [25]. Other inflammatory and tissue injury biomarkers have also been widely studied in COVID-19. Lactate dehydrogenase is an intracellular enzyme released during cellular injury, and elevated serum levels reflect tissue damage, particularly in the lungs, where severe infection and hypoxia may lead to cell destruction and increased leakage of this enzyme into the circulation [12]. Ferritin, an intracellular iron storage protein, is another important marker because hyperferritinemia reflects both iron dysregulation and an intense inflammatory state, and high ferritin levels have been linked to severe disease, cytokine storm, and increased mortality [13]. Similarly, the neutrophil-to-lymphocyte ratio has emerged as a simple and useful indicator of inflammation and disease severity, as severe COVID-19 is often associated with neutrophilia and lymphopenia, reflecting immune dysregulation and poor clinical outcome [14]. Pregnancy represents a special physiological state in which significant immunological, respiratory, cardiovascular, and hematological adaptations occur, making pregnant women potentially more vulnerable to infectious diseases, including COVID-19 [29]. During pregnancy, the maternal immune system undergoes dynamic modulation to tolerate the semi-allogeneic fetus while maintaining defense against pathogens, and this balance may be disturbed by SARS-CoV-2 infection, especially during the naturally pro-inflammatory first and third trimesters [1,2]. COVID-19 in pregnancy has been associated with increased risks of maternal complications such as severe pneumonia, preeclampsia, preterm labor, cesarean delivery, and adverse neonatal outcomes, although the exact magnitude of these risks continues to be investigated. Because interleukin-6 is a central mediator in the inflammatory cascade of COVID-19 and may contribute to both maternal and fetal complications, assessing its serum level in pregnant women infected with COVID-19 may provide valuable insight into the degree of inflammation, disease severity, and possible obstetric consequences [12]. The aim of this study was to evaluate the role of interleukin-6 serum level in inflammation in covid-19 pregnant women.

A prospective observational study was conducted in Tikrit city from the 1st of January to the end of September 2022 to evaluate inflammatory biomarkers among pregnant women with and without COVID-19 infection. The study was carried out at the Department of Obstetrics and Gynecology in Salah Al-Din General Hospital, where all enrolled participants were followed until delivery. A total of 140 pregnant women were included and equally divided into two groups: Group A consisted of 70 pregnant women diagnosed with COVID-19 infection, and Group B included 70 apparently healthy pregnant women without any known medical disorders. The diagnosis and classification of COVID-19 severity were determined by specialized physicians in internal medicine according to national guidelines and international standards. Patients were considered suspected cases if they presented with one or more of the following clinical features: fever greater than 37.8°C, cough, dyspnea, loss of taste or smell, or diarrhea. The clinical spectrum ranged from asymptomatic infection to severe disease requiring hospitalization and oxygen therapy, including cases progressing to acute respiratory distress syndrome, based on the classification of the China Center for Disease Control and Prevention.

Strict exclusion criteria were applied to minimize confounding factors. Pregnant women with multiple pregnancies, those with pre-existing maternal medical conditions such as diabetes mellitus, hypertension, or chronic systemic diseases, and those with a history of preterm delivery or stillbirth in previous pregnancies were excluded from the study. Ethical approval was obtained from the Council of the College of Medicine, Tikrit University, and official permission was granted by the Salah Al-Din Health Directorate and the administration of Salah Al-Din General Hospital. All participants were informed about the objectives of the study, and informed consent was obtained prior to enrollment. Data were collected through direct interviews using a structured questionnaire designed by the investigator, which included demographic data such as age, weight, height, and relevant clinical and obstetric history.

Venous blood samples were collected from all participants under aseptic conditions. Approximately five milliliters of blood were drawn from each subject without the use of a tourniquet to avoid hemoconcentration. The samples were divided into two portions: two milliliters were transferred into tubes containing sodium citrate as an anticoagulant for complete blood count analysis using an automated hematology analyzer, while the remaining three milliliters were placed into gel tubes for serum separation. After clot formation, samples were centrifuged at 3000 rpm for 15 minutes, followed by a second centrifugation step for 10 minutes to ensure clear serum separation. The obtained serum was aliquoted into labeled tubes and stored at −20°C until biochemical analysis.

Serum interleukin-6 (IL-6) levels were determined using a sandwich enzyme-linked immunosorbent assay (ELISA) technique. Microtiter plates pre-coated with specific antibodies were used to capture IL-6 from the samples. Following incubation with biotin-conjugated antibodies and enzyme-linked avidin, a substrate solution was added, producing a colorimetric reaction proportional to the IL-6 concentration. The reaction was terminated using sulfuric acid, and absorbance was measured at 450 nm using an ELISA reader. IL-6 concentrations were calculated from a standard curve, with normal values considered less than 6.6 pg/mL.

Serum ferritin and C-reactive protein (CRP) levels were measured using immunofluorescence-based assays with the ichroma™ system. These assays relied on antigen-antibody interactions producing fluorescence signals proportional to analyte concentration, which were automatically calculated and displayed by the analyzer. Ferritin levels were expressed in ng/mL, with normal reference values ranging from 20 to 250 ng/mL for women, while CRP levels were expressed in mg/dL with normal values up to 6 mg/dL. Lactate dehydrogenase (LDH) activity was measured using an enzymatic method based on the conversion of lactate to pyruvate, with the rate of NADH formation measured photometrically and directly proportional to enzyme activity. Normal LDH values ranged from 135 to 225 U/L. Complete blood count parameters were analyzed using an automated hematology analyzer to assess leukocyte and lymphocyte counts.

All laboratory kits used in the study were obtained from standardized commercial sources, including MYBIOSOURCE (USA) for IL-6 ELISA kits, Roche Cobas (Germany) for LDH analysis, Swelab (Switzerland) for complete blood count, and ichroma™ (Korea) for ferritin and CRP assays. The instruments used included ELISA washers and readers (Biotek, USA), centrifuges (Presvac, Argentina), refrigerators (Toshiba, Japan), and the ichroma™ analyzer system.

Statistical analysis was performed using the Statistical Package for Social Sciences (SPSS) version 23.1. Quantitative variables were expressed as mean ± standard deviation, while qualitative variables were presented as frequencies and percentages. Comparisons between groups were conducted using the independent t-test for continuous variables and chi-square test for categorical variables. The level of statistical significance was determined based on p-values, where values greater than 0.05 were considered non-significant, values between 0.01 and 0.05 were considered statistically significant, and values less than 0.01 were considered highly significant.

In comparative analysis of demographic characteristics between pregnant women who have COVID-19 infection and pregnant women who do not have any known risk factors for the infection, the study showed no significant differences between the studied groups regarding maternal age, parity, BMI and gestational age at hospital admission.

The study showed that, the highest mean levels of IL-6 was found in pregnant women with COVID-19 infection (18.5±2.12pg/ml) and the lowest mean was in Healthy Pregnant women (3.58±1.28 pg/ml), (P:0.001). The study also showed that WBC, Neutrophil to lymphocyte ratio, serum Ferritin and LDH significantly elevated in Pregnant women with COVID-19 infection while mean platelets count was reduced significantly in Pregnant women with COVID-19 infection, Table 2.

Table 1: Demographic features of studied groups

Table 2: Comparative analysis of IL-6 and other Lab parameters in pregnant with and without COVID-19 infection

The study demonstrated that 67.14% of Pregnant women with COVID-19 were with mild infection, 24.29% with moderate infection and 8.57% were with Severe and Critical infection, Table 3.

Table 3: Distribution of Pregnant women with COVID-19 infection according to severity of the disease

The study demonstrated that the rate of Pregnancy complication was 18.57% (13 of 70) of pregnant women with COVID-19 infection compared with 1.43% of healthy pregnant women (P-value:0.001) Table 4.

Table 4: Pregnancy complication rate in pregnant with and without COVID-19 infection

P-value:0.001

The study demonstrated that high rate of women with pregnancy complication (33.33%) were with preterm delivery,  26.67%  with  preeclampsia, 13.33%   with   low birth weight /Still birth and 6.67% with each of gestational diabetes and missed abortion, Table 6 and Figure 1.

Table 6: Distribution of Pregnancy complication 

The study demonstrated that most of women with pregnancy complications (preterm delivery, preeclampsia, and missed abortion) were recorded among pregnant women with severe COVID-19 infection, Table 7.

Table 7: Distribution of Pregnancy complication according to severity of COVID-19 infection

P-value 0.001

Figure 1: Pregnancy complication rate in pregnant with and without COVID-19 infection

Figure 2: Distribution of Pregnancy complication

The study showed that, of IL-6 was significantly elevated in pregnant women with COVID-19 infection and Pregnancy complication (19.21±2.45 pg/ml) and the lowest mean was in without Pregnancy complication (16.48±1.92 pg/ml), (P:0.021). The study also showed that  Neutrophil to  lymphocyte ratio,  serum  Ferritin  and 

Table 8: Relation of IL-6 and other Lab parameters in Pregnancy complication of in Covid-19 infected women  LDH significantly elevated in Pregnant women with COVID-19 infection and Pregnancy complication infection, Table 8.

The main findings of the present study indicated that pregnancy complications and inflammatory markers were significantly higher in pregnant women with COVID-19 infection. Furthermore, significantly higher levels of interleukin-6 (IL-6), ferritin, neutrophil-to-lymphocyte ratio (NLR), and lactate dehydrogenase (LDH), together with significantly lower platelet counts, were observed in pregnant women with COVID-19 infection. Additionally, cytokine levels were significantly correlated with disease severity. Pregnancy is associated with a unique immune adaptive process that allows proper implantation of the semi-allogeneic fetus [1]. Increased production of pro-inflammatory cytokines such as IL-6 has been linked to higher rates of pregnancy complications including miscarriage and preterm delivery; however, these inflammatory processes are also essential for physiological events such as placental invasion and parturition [2]. Therefore, maintaining a balance between pro-inflammatory and anti-inflammatory cytokines is critical for a healthy pregnancy [3]. The excessive inflammatory response observed in COVID-19 infection is considered a major pathophysiological mechanism contributing to morbidity and mortality [4]. Dysregulated cytokine production, particularly IL-6, has been reported in severe cases and is associated with poor clinical outcomes [5]. Consequently, recent therapeutic approaches have focused on modulating immune responses in COVID-19 patients [6]. However, pregnancy represents a complex immunological state, making it difficult to predict the safety and efficacy of immunomodulatory therapies in this population [7,8]. In agreement with our findings, Tanacan et al. [9] reported that pregnant women with SARS-CoV-2 infection had significantly higher rates of pregnancy complications, as well as elevated levels of ferritin, LDH, NLR, and IL-6 (p<0.05). The significantly higher IL-6 levels observed in the present study, particularly among women with pregnancy complications, may be explained by the shift toward a pro-inflammatory state in severe COVID-19 cases. Similarly, Monteleone et al. [6] demonstrated that IL-6 is associated with adverse pregnancy outcomes such as preterm delivery, premature rupture of membranes, and chorioamnionitis. Elevated IL-6 is also recognized as a key mediator in the cytokine storm characteristic of severe COVID-19 [4]. The absence of significant variation in IL-6 levels across pregnancy trimesters in our study is consistent with previous findings. However, our results differ from those reported by Jamilloux et al. [5], who did not find a significant association between IL-6 levels and pregnancy complications in COVID-19 cases. In the present study, white blood cell count and NLR were significantly elevated, while platelet counts were significantly reduced in pregnant women with COVID-19 infection. These findings are consistent with Carranza et al. [10], who reported elevated NLR levels in pregnant women with COVID-19, particularly in those with complications, suggesting that NLR reflects systemic inflammatory responses. The differentiation of leukocyte subtypes represents an immune response to inflammation, leading to changes in NLR values across various inflammatory conditions. Significant differences in leukocyte counts and NLR between healthy and infected pregnant women have also been reported, with higher values associated with disease severity [11]. Our findings are further supported by Lombardi et al. [12], who highlighted that COVID-19 is characterized by a pronounced systemic inflammatory response contributing to morbidity and mortality. Elevated NLR, LDH, and C-reactive protein (CRP) levels were associated with disease severity, although their study did not observe a significant increase in total white blood cell count. Several studies have suggested that severe COVID-19 infection may increase the risk of adverse pregnancy outcomes [13-18]. Additionally, emerging evidence indicates an association between the pandemic and complications such as gestational hypertension, fetal growth restriction, and preeclampsia [19-21]. The increased risk of adverse outcomes may be attributed to pregnancy-induced immune modulation, which in some cases leads to increased susceptibility to infections, while in others may provide partial protection [22,23].

The study demonstrated that most of women with pregnancy complications (preterm delivery, preeclampsia, and missed abortion) were recorded among pregnant women with severe COVID-19 infection and interleukin-6, Neutrophil to lymphocyte ratio, serum Ferritin and lactate dehydrogenase significantly elevated in pregnant women with COVID-19 infection and Pregnancy complication and the low mean was in without Pregnancy complication.

1. Robertson, S. A. “Immune Regulation of Conception and Embryo Implantation.” Journal of Reproductive Immunology, vol. 85, 2010, pp. 51–57.

2. Chow, S. S., et al. “Cytokine Levels in Pregnancy.” Cytokine, vol. 44, 2008, pp. 78–84.

3. Raghupathy, R., and J. Kalinka. “Cytokine Imbalance in Pregnancy Complications.” Frontiers in Bioscience, vol. 13, 2008, pp. 985–994.

4. Castelli, V., A. Cimini, and C. Ferri. “Cytokine Storm in COVID-19.” Frontiers in Immunology, vol. 11, 2020, p. 2132.

5. Jamilloux, Y., et al. “Cytokine Interventions in COVID-19.” Autoimmunity Reviews, vol. 19, 2020, p. 102567.

6. Monteleone, G., P. Sarzi-Puttini, and S. Ardizzone. “Anticytokine Therapy in COVID-19.” The Lancet Rheumatology, vol. 2, 2020, pp. e255–e256.

7. Azizieh, F., et al. “Cytokine Profiles in Pregnancy.” American Journal of Reproductive Immunology, vol. 79, 2018, p. e12818.

8. PrabhuDas, M., et al. “Immune Mechanisms in Pregnancy.” Nature Immunology, vol. 16, 2015, pp. 328–334.

9. Tanacan, A., et al. “Cytokine Profile in Pregnant COVID-19 Patients.” Cytokine, vol. 140, 2021, p. 155431.

10. Carranza-Lira, S., and M. Garcia-Espinosa. “NLR in Pregnant Women with COVID-19.” International Journal of Gynecology & Obstetrics, vol. 157, 2022, pp. 296–302.

11. Fathi, N., and N. Rezaei. “Lymphopenia in COVID-19.” Cell Biology International, vol. 44, 2020, pp. 1792–1797.

12. Lombardi, A., et al. “Inflammatory Biomarkers in Pregnancy COVID-19.” Scientific Reports, vol. 11, 2021, p. 13350.

13. Delahoy, M. J., et al. “COVID-19 Outcomes in Pregnancy.” MMWR, vol. 69, 2020, pp. 1347–1354.

14. WAPM Working Group. “Maternal Outcomes COVID-19.” Ultrasound in Obstetrics & Gynecology, vol. 57, 2021, pp. 232–241.

15. Khoury, R., et al. “Pregnancy Outcomes COVID-19.” Obstetrics & Gynecology, vol. 136, 2020, pp. 273–282.

16. London, V., et al. “COVID-19 Pregnancy Outcomes.” American Journal of Perinatology, vol. 37, 2020, pp. 991–994.

17. Prabhu, M., et al. “SARS-CoV-2 Pregnancy Cohort.” BJOG, vol. 127, 2020, pp. 1548–1556.

18. Sakowicz, A., et al. “Risk Factors COVID-19 Pregnancy.” AJOG MFM, vol. 2, 2020, p. 100198.

19. Stowe, J., et al. “Stillbirth during COVID-19.” JAMA, vol. 325, 2021, pp. 86–87.

20. Khalil, A., et al. “Preterm Delivery COVID-19.” JAMA, vol. 324, 2020, pp. 705–706.

21. Handley, S. C., et al. “Birth Outcomes COVID-19.” JAMA, vol. 325, 2021, pp. 87–89.

22. Silasi, M., et al. “Viral Infections in Pregnancy.” American Journal of Reproductive Immunology, vol. 73, 2015, pp. 199–213.

23. Mor, G., and I. Cardenas. “Immune System in Pregnancy.” American Journal of Reproductive Immunology, vol. 63, 2010, pp. 425–433.