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 Table of Contents  
ORIGINAL ARTICLE
Year : 2021  |  Volume : 5  |  Issue : 1  |  Page : 69-73

Serum levels of interleukin-6, ferritin, C-reactive protein, lactate dehydrogenase, D-dimer, and count of lymphocytes and neutrophils in COVID-19 patients: Its correlation to the disease severity


1 Department of Science, College of Basic Education, University of Waist, Al-Kut City, Iraq
2 Department of Nursing, Amara Technical Institute, Southern Technical University, Amara City, Iraq

Date of Submission29-Oct-2020
Date of Acceptance24-Dec-2020
Date of Web Publication13-Mar-2021

Correspondence Address:
Dr. Younus Jasim Abdullah
Department of Medical Laboratory Technology, Amara Technical Institute, Al-Kahlaa Street, Amara City, Maysan Province
Iraq
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/bbrj.bbrj_188_20

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  Abstract 


Background: Since its first emergence in Wuhan city, China, severe acute respiratory syndrome coronavirus 2, which is responsible for the pandemic COVID-19, has become a significant health problem all over the world affecting over 2.1 million people globally. Methods: The current study aimed to investigate serum levels of interleukin 6 (IL-6), ferritin, C-reactive protein (CRP), lactate dehydrogenase (LDH), D-dimer as well as lymphocytes and neutrophils count in COVID-19 patients, and to clarify the correlation of these parameters with disease severity and progression. For these purposes, (100) patients with COVID-19 (confirmed by polymerase chain reaction) and (20) apparently healthy people (with matched age and sex) were included in the current study and considered as a control group. Results: All study population (patients and control) were subjected to the evaluation of serum levels of IL-6, ferritin, CRP, LDH, D-dimer, as well as lymphocytes and neutrophils, count. COVID-19 patients showed a significant elevation in the levels of all parameters included in this study when compared with healthy controls. We also found that all of IL-6, ferritin, CRP, LDH, D-dimer are significantly associated with the severity of the COVID-19 symptoms. Conclusion: Lymphopenia and increased neutrophils were also effectively correlated with disease progression. In line with these results, we concluded a proportional correlation between the aforementioned parameters and COVID-19 suggesting the uses of these tests to the diagnosis of critical cases.

Keywords: COVID-19, interleukin 6, C-reactive protein, D-dimer, ferritin, lymphocyte, neutrophils


How to cite this article:
Kadhim AS, Abdullah YJ. Serum levels of interleukin-6, ferritin, C-reactive protein, lactate dehydrogenase, D-dimer, and count of lymphocytes and neutrophils in COVID-19 patients: Its correlation to the disease severity. Biomed Biotechnol Res J 2021;5:69-73

How to cite this URL:
Kadhim AS, Abdullah YJ. Serum levels of interleukin-6, ferritin, C-reactive protein, lactate dehydrogenase, D-dimer, and count of lymphocytes and neutrophils in COVID-19 patients: Its correlation to the disease severity. Biomed Biotechnol Res J [serial online] 2021 [cited 2021 Dec 3];5:69-73. Available from: https://www.bmbtrj.org/text.asp?2021/5/1/69/311091




  Introduction Top


Since its first emergence in Wuhan city, China, at the end of 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a positive sense single-stranded enveloped virus which is responsible for the pandemic COVID-19 has become a significant health problem all over the world with over 2.1 million cases and 120,000 deaths so far.[1] In Iraq, the first case of COVID-19 was recorded in March 2020 in Al Najaf city, since which the virus spread dramatically around the country causing approximately 51,300 infections and more than 7000 deaths according to the daily reports declared by the Iraqi ministry of health.

The clinical spectrum of SARS-CoV-2 infection appears to be wide, encompassing asymptomatic infection, mild upper respiratory tract illness, and severe viral pneumonia with respiratory failure and even death, with many patients being hospitalized with pneumonia.[2]

Urgent identification of clinical and laboratory predictors of progression to serious and lethal forms is urgently needed in the battle against COVID-19 disease. These predictors will allow clinical management, direct inductive research approach to target patients at increased risk of developing a serious disease and optimize the allocation in the ongoing pandemic of limited human and technological resources. Besides, the identification of laboratory criteria capable of distinguishing between severe and nonserious cases, or those at high or low mortality risk, would enable the enhancement of clinical understanding of the situation.[3]

There is overwhelming evidence that there are features of hyper inflammation in critically ill patients, consisting of elevated serum C-reactive protein (CRP), procalcitonin, D-Dimer, and hyperferritinemia.[4] Furthermore, lymphopenia is strongly correlated with the seriousness of the disease. Patients who died from COVID-19 have substantially lower lymphocyte counts than survivors. Other blood cells, such as neutrophils, eosinophils, platelets, as well as CD8+ cell counts, have been partially considered as determinants in discriminating mild from severe COVID-19, the significance of which is still vague.[5] Cytokine storms may also be related to disease severity. Increased serum expression of interleukin (IL)-2R and IL-6 appears to be predicting the severity and prognosis of COVID-19 patients. Pathological analysis of a biopsy sample from a patient who died from COVID-19 showed interstitial mononuclear inflammatory infiltration in both lungs. Furthermore, the peripheral cytometric examination of blood flow revealed that overactivation of T-cells compensated, in part, for the extreme immune injury in this patient. These results indicate a potentially important role in the pathophysiology of the cytokine storm COVID-19.[6]

In line with the aforementioned, this study aimed to estimate serum levels of IL-6, D dimer, ferritin, CRP as well as total blood lymphocyte counts in patients with COVID-19 disease, in addition, to explore the correlation between the levels of the above parameters and severity of the disease. To the best of our knowledge, this is the first study in Iraq that considers such goals in patients infected with SARS-CoV-2.


  Methods Top


Study population

The current study involved (100) patients suffering from COVID-19, aging between (18 and 45) years old and of both sexes. In addition, (50) apparently healthy persons of the same ages and sexes were included in this study as a control group. COVID-19 patients were grouped into two groups (simple to moderate and severe) according to their symptom's severity (fever, cough, respiratory distress, loss of sense of taste and smell, and needing for ventilation). All of the patients and healthy persons were subjected to the estimation of serum IL-6, D-Dimer, lactate dehydrogenase (LDH), CRP, ferritin, as well as the total count of lymphocyte and neutrophils.

Study sampling

Five milliliters of venous blood is collected from the study groups and divided into two tubes (plane tubes and Ethylenediaminetetraacetic acid [EDTA] tubes). The first part of the blood samples (EDTA tubes) is applied to the hematology autoanalyzer (Pentra 80. manufactured by the ABX-Horiba group, Minami-Ku Kyoto Japan). The second part of the blood samples was centrifuged to obtain serum which is used for the determination of serum concentration of IL-6, D-Dimer, LDH, CRP, and ferritin.

Estimation of serum interleukin-6

Electrochemiluminescence immunoassay technique and (Cobas) instrument are used for the detection of serum IL-6 in patients and the control group using a kit performed by Roche company (Roche Diagnostics GmbH, Sandhofer Strasse 116, D-68305 Mannheim. 2020).

Determination of serum D-dimer and C-reactive protein

Serum concentrations of D-Dimer and CRP were evaluated using a specific automated protein analyzer (PA120) provided by (Shenzhen Genius Electronics Co., Ltd. China 2019). Serum samples for each of the patients and healthy persons were applied to the instrument then the concentrations of D-dimer and CRP are calculated automatically.

Total leukocytes and neutrophils counting

Anticoagulated blood samples from both patients and the control group are applied to a hematology autoanalyzer (Pentra 80. manufactured by ABX-Horiba group, Minami-Ku Kyoto Japan) to estimate total counts of white blood cells (WBCs) and neutrophils. Samples are processed by the instrument then total WBCs and neutrophils are calculated automatically.

Statistics

The results of the current study are analyzed using (SPSS Statistics 20; 1 New Orchard Road Armonk, New York 10504-1722 USA) to find the variation in the results of the study. Statistical significance was defined as a (P < 0.05). Statistical tests such as (independent t-test, percentage) were used for data analysis.


  Results Top


The results of the current study, which included (100) patients suffering from SARS CoV2 infection, found, as illustrated in [Figure 1], that the percent of COVID-19 infection was higher in women (60%) than men (40%).
Figure 1: Percent of COVID-19 infection according to sex

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The results of [Table 1] are showing that the mean of the serum level of IL-6 was higher significantly (P ≤ 0.000) in COVID-19 patients (26.72 ± 15.05 pg/ml) compared to the control group (5.38 ± 0.89 pg/ml). Likewise, the mean of the serum levels of CRP, ferritin, LDH, and D-Dimer was also elevated significantly among COVID-19 patients compared to the control group.
Table 1: The differences in the serum levels of interleukin-6, ferritin, lactate dehydrogenase, C-reactive protein and D-Dimer in COVID-19 patients and control group

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We also found that total lymphocyte count was decreased significantly (P = 0.018) in patients infected with SARS CoV2 (mean = 13.66 ± 4.29), in contrast, neutrophils count was increased significantly (P ≤ 0.000) in patients (90.43 ± 12.40) compared with healthy subjects (51.63 ± 5.66), [Table 2].
Table 2: The total count of lymphocytes and neutrophils in COVID-19 patients and control group

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The results of the current study revealed, as shown in [Table 3], that all of IL-6, ferritin, LDH, and CRP, except D-Dimer, are associated with severity of SARS CoV2 infection. The mean of theses parameters is significantly increased in patients with severe coronavirus infection and reached (28.63 ± 14.67 pg/ml, 471.60 ± 202.92 μg/ml, 119.98 ± 68.54 mg/ml, and 443.55 ± 99.38 IU/L) for IL-6, ferritin, rapid respiratory panel( RRP), and Lactate Dehydrogenase(LDH) respectively, while in patients with simple to moderate infection, the levels of IL-6, ferritin, CRP, and LDH is reached (9.56 ± 0.66 pg/ml, 232.14 ± 161.29 μg/ml, 25.64 ± 22.53 mg/ml, and 335.80 ± 97.63 IU/L) respectively. Furthermore, the D-Dimer level was found to decreased non significantly (P = 060) in a patient with severe infection (1.68 ± 0.30) compared to a patient with mild-to-moderate symptoms [Table 3].
Table 3: Differences in the levels of interleukin-6, ferritin, lactate dehydrogenase, C-reactive protein, and D-Dimer in COVID-19 patients according to the severity of infection

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In the same context, we found, as shown in the [Table 4], that lymphocytes are decreased significantly (P = 0.012) in patients with severe COVID-19 disease, while neutrophils are significantly increased (P = 0.013) in patients with severe symptoms of SARS CoV2 infection.
Table 4: Differences in lymphocytes and neutrophils count in COVID-19 patients according to the severity of infection

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  Discussion Top


In the present study, which is conducted in Baghdad city, 60% of the SARS CoV2-infected patients were women and 40% were men. In agreement with our results, Gebhard et al., found that female's percentage of infection were higher than males and reached (52%) in Switzerland and Belgium, (54%) in Portugal, (53%) in France, and (60%) in South Korea.[7]

Females and males have a variable response to viral infection just such as SARS CoV, Middle East respiratory syndrome (MERS) CoV, SARS CoV2, and other viruses. These differences are the leading of disease severity and incidence between the two genders. Multiple factors contribute to the disparity in sex-specific disease outcomes following virus infections. Sex-specific steroids and the activity of X-linked genes, both of which modulate the innate and adaptive immune response to virus infection, influence the immune response. Furthermore, the differences in the expression of angiotensin-converting enzyme (ACE) 2 receptor and the cellular serine protease TMPRSS2, which are necessary for the binding and priming of SARS CoV2, may have an important role.[7],[8] We also believe that the increased stress and domestic work during the pandemic especially in Iraq may also affect the capabilities of the female immune system against COVID-19.

There is a significant increase in the serum levels of IL-6, ferritin, CRP, LDH, and D-Dimer in COVID-19 patients [Table 1]. In addition, the levels of theses parameters are effectively associated with the diseases severity in the patients as shown in [Table 3]. Zhu et al., 2020[9] reported that IL-6 is elevated in COVID-19 patients and reached to (24.11 pg/ml) and its level was significantly correlated with the severity of the symptoms in COVID-19 patients. Similarly, Huang et al., in 2020[10] found that IL-6 is increased in patients with SARS CoV2 infection and its high level was closely associated with the level of virus RNA in the blood and disease progression. It is stated that during viral infection, IL-6 together with transforming growth factor-beta induces the differentiation of naïve CD4 into Th17 cells, which are important for the defense against viruses and other pathogens at mucosal sites. In addition, there is synergic interaction between IL-6 and IL-7 and IL-15 to induce the differentiation and catalytic ability of CD8 T cells which is important in the response against viral infections.[11]

The ferritin level in COVID-19 patients, as shown in Table 1, was significantly increased and reached to (447.65 ± 210.62 μg/mL). Likewise, the serum level of CRP was also elevated significantly in patients when compared with the healthy group. The elevation of ferritin and CRP was correlated with the disease progression [Table 3]. Several publications have shown that elevated ferritin levels have been associated with worse outcomes, along with several other pro-inflammatory markers, involving CRP and IL-6, and may even help predict these outcomes.[12],[13],[14] Elevated ferritin levels due to secondary hemophagocytic lymphohistiocytosis and cytokine storm syndrome have been reported in severe COVID-19 patients. It has been proposed that COVID-19 could be part of the broader spectrum of hyperinflammatory syndromes such as the secondary hemophagocytic lymphohistiocytosis.[15] Hyperferritinemia is a cardinal characteristic of these syndromes. COVID-19 severity and worse prognosis mean that mortality could be due to virally induced hyper inflammation which is substantially associated with high ferritin level. However, the circulating levels of ferritin can not only represent a strong phase response but play an important role in inflammation.[16] In COVID-19 patients, CRP levels were increased and it was shown that survivors had a median CRP of approximately 40 mg/L, while nonsurvivors had a median CRP of (125 mg/L), suggesting a good association with the severity and prognosis of the disease.[5]

In addition to IL-6, ferritin, and CRP, our results also found a significant elevation in the level of LDH as well as D-Dimer in COVID-19 patients. However, the increases in both D-Dimer and LDH were effectively correlated with disease severity as shown in Table 3. D-Dimer is a commonly used fibrin degradation test used to diagnose thrombotic diseases at an early level.[17] Prior studies have shown that D-Dimer level is higher in extreme cases of community-acquired pneumonia and chronic obstructive pulmonary disease and can be used as a significant biomarker and that D-Dimer >1 μg/mL is one of the mortality risks factors in adults with COVID-19 who receive the medication for their pneumonia in a group acquired environment.[18],[19] It was also stated that D-Dimer at admission more than 2.0 μg/mL (fourfold increase) could effectively be correlated with disease severity and mortality in COVID-19 patients, as shown in Table 3 in this review. These results indicated that D-Dimer could be an early and helpful marker to improve the management of COVID-19 patients.[17],[20],[21]

LDH is a glycolytic cytoplasmic enzyme present in virtually every tissue. In general, its elevation suggests tissue injury. In patients infected with MERS-CoV, increased LDH was a typical finding in.[22],[23] Independent mortality factors have been identified for patients with the extreme acute respiratory syndrome[24] and H1N1 infection.[25] Possible subclinical tissue damage was indicated by our observation of increased LDH in the early stage of extreme COVID-19 cases. Although the virus binds to the human ACE2 receptor in the lungs,[26],[27] which explains why the lungs are the first affected organs, different cytokine abnormalities and multiple organ dysfunction can be observed in serious patients as the disease progresses, suggesting systemic organ damage caused by excessive immune system activation.[28],[29]

Results of Table 2 showed that COVID-19 patients have significantly decreased lymphocyte count (P = 0.018) and significantly increased neutrophils count (P ≤ 0.000). The decreased and elevated lymphocyte and neutrophils count was significantly correlated to the disease severity as shown in the Table 4.

Hematologic variations were established as supporting evidence of infection with COVID-19 and as potential signs of serious illness. Numerous clinical guidelines define that potential SARS-CoV-2 infection, especially in severe cases, exhibits irregularities in hemocytometry. The U. S. Centers for Disease Control and Prevention have issued guidelines emphasizing that the most common laboratory abnormalities recorded in hospitalized COVID-19 patients with pneumonia were leukopenia (92%–5%), leukocytosis (24%–30%), and lymphopenia (63%).[30]

Lymphopenia was also a clear finding and experiments using flow cytometry showed that these improvements were correlated with lower counts of CD4+ and CD8+ T lymphocytes[31],[32] In the same context, Song et al. 2020[33] have revealed that the most consistent abnormal hemocytometric findings in COVID-19 patients are lymphopenia and increased neutrophils, and these changes can also increase throughout the disease, especially in those with serious illness. In pregnant women with COVID-19, lymphopenia was also reported; however, this finding is even less significant in infected children.[34]


  Conclusion Top


Contrary to previous studies, our current study found that male sex may be at low risk to be infected with SARS CoV2. Infection with COVID-19 has led to significant elevation in the levels of IL-6, ferritin, CRP, LDH, D-dimer as well as lymphopenia and increased neutrophils count. These differences were effectively associated with disease severity and progression, suggesting the use of such clinical markers to recognize severe illnesses.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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