In ammatory Cytokine Levels in Pulmonary Embolism Severity: A Retrospective Study

Xiaoming Li Xinhua Hospital A liated to Shanghai Jiaotong University School of Medicine Yukun Li Dezhou people’s hospital Wenjing Ye Xinhua Hospital A liated to Shanghai Jiaotong University School of Medicine Xi Chen Xinhua Hospital A liated to Shanghai Jiaotong University School of Medicine Xuejun Guo Xinhua Hospital A liated to Shanghai Jiaotong University School of Medicine Wen Gu (  guwen@xinhuamed.com.cn ) Xinhua Hospital A liated to Shanghai Jiaotong University School of Medicine


Introduction
Venous thromboembolism (VTE), clinically presenting as deep vein thrombosis (DVT) or PE, is a common disorder with an estimated incidence of one to three per 1,000 population annually [1]. A rising tendency in VTE incidence rates has been revealed in epidemiological studies [2]. As reported, the incidence of VTE currently is almost 10-30 times higher than that in 20 years ago in China. Pulmonary embolism (PE) as the most frequent acute syndrome in respirology department is a potentially life-threatening disease.
Accurate and prompt diagnosis is essential for reducing mortality. Once PE is diagnosed, risk strati cation should be performed immediately. PE severity can be classi ed to low risk, intermediate risk  [3].
Increasing evidence revealed that cytokines are involved in the development of VTE and may in uence the mortality [4]. These new insights into the role of cytokines in VTE have aroused Clinicians' attention to anti-in ammatory therapy [5]. A recent review summarized the interplay between cytokines and venous thrombosis. Cytokines including IFNγ, IL-6, IL-17A, IL-9, IL-1β and TGF-β perform pro-thrombotic activity, while other cytokines such as IL-10, TNF-α and IL-8 appear to promote thrombus resolution in late phase of venous thromboembolism [6]. Anomalous cytokines levels and high PE frequency were found in patients with cancer, pneumonia, tuberculous pleural effusion, and other disease [7][8][9]. Abnormal cytokine levels caused by primary disease may involve in the development of PE. But the relationship between cytokines and PE severity is unclear. Whether comorbidities would in uence PE severity through cytokines is worth studying.
In the present study, we evaluated the relationship between cytokines and PE severity. in addition, we investigated the association of comorbidities and the severity of PE in patients.

Study design and participants
A retrospective study was conducted to investigate the association between cytokines as well as complications and PE severity. All the patients with con rmed diagnosis of PE in Xinhua hospital a liated to Shanghai Jiao Tong University School of Medicine between Jan, 2019 and Apr, 2021 were reviewed. Interleukin (IL)-8, IL-6, IL-2R, tumor necrosis factor (TNF-a) and C-reactive protein (CRP), white blood cell (WBC) counts, erythrocyte sedimentation rate (ESR) as well as the rate of comorbidities were compared between different PE severity groups. The association between cytokines, in ammatory makers, comorbidities, and PE severity was analyzed.
This retrospective study only collected the clinical data of patients, did not interfere with the treatment plan of patients, and did not pose any risk to patients' physiology. Due to the retrospective nature of the study, informed consent was waived. All protocols were approved by the Ethics Committee of Xin Hua

Data Collection
In our hospital, PE severity was strati ed as soon as possible according to clinical appearance, systolic blood pressure, imaging for right ventricular dysfunction (CTA or echocardiography) and/or biomarkers (troponin, NT-pro-BNP) as described before [10]. According to the medical record, clinical data includes demographic information (age, gender, previous medical history, comorbidities), PE severity, laboratory tests (CRP, WBC counts, ESR, D-dimer, IL-6, IL-8, TNF-α, and IL-2 receptor) was collected.
Statistical analysis SPSS version 22.0 (IBM Corporation, New York) was used for the statistical analysis. Data were presented as median with interquartile range. Categorical data were presented as frequencies or percentages.
Comparison of median among groups was performed by using Kruskal-Wallis test. Comparison of rates among groups was performed by Chi-square test. The association between cytokines and the severities of PE was analyzed by using Spearman rank-order correlation analysis. The association between commodities and the severities of PE was analyzed by using ordinal multinomial Logistic regression analysis. Statistical signi cance was set at P < 0.05.

Results
The demographics and clinical characteristics of patients.
Correlation between biomarkers and PE severity.

Discussion
In the present study, we investigated the relationship between plasma cytokines as well as comorbidities and PE severity. To the best of our knowledge, this is the rst study to investigate the association between cytokines and PE severity in the literature. According to our study, a negative correlation between IL-8 levels and PE severity was found. A weak negative correlation between TNF-α and PE severity was also found. We didn't observe any signi cant correlation between cytokines like IL-6, IL2R, CRP, WBC, ESR, and PE severity. In addition, we found that patients with comorbidities regarding coronary heart disease, deep venous thrombosis, and respiratory failure may have higher risk to surfer a more sever PE.
IL-8, the most important C-X-C chemokine, is produced by both in ammatory cells and structural cells including monocytes, neutrophils, endothelial cells, and epithelial cells. It was the most powerful chemoattractant of neutrophils [11]. As previous study, elevated IL-8 level was found in PE patients compared to control [12]. According to our study, once regarding to PE severity, elevated IL-8 may be a protect factor. This result was consistent with Henke's study. Used rats VTE model, Henke and colleagues found that extrinsic IL-8 treatment enhances thrombus resolution through neovascularization and neutrophil recruitment. Invaded neutrophils modulate collagen and brin degradation through the secretion of matrix metalloproteinases (MMP) in the thrombus. In addition, neovascularization is part of thrombus remodeling and involves the vascular endothelial growth factor (VEGF) that contributes to vein recanalization [13]. Whether the decreased level of IL-8 contributes to the severity of pulmonary embolism by affecting thrombus absorption needs further research. Nosaka and colleagues found that TNFα may have a bene cial role during thrombus resolution by promoting brinolysis, collagenolysis and neovascularization [14]. In our study, the negative correlation between TNFα and PE severity is week. Whether a strong correlation could occur when increase the sample size is need to further research.
Elevated IL-6 circulating levels were found in VTE patients compared to healthy controls which indicated an important role of IL-6 in VTE even the underlying mechanism is unclear [15,16]. But in our study, there was no signi cant difference of IL-6 among groups and no signi cant correlation between IL-6 and PE severity.
Further, we investigated whether the presence of comorbidity at the time of diagnosis of PE could in uence PE severity. As our data shown, PE patients with coronary heart disease, DVT, and respiratory failure had increased risk of 3.76-fold, 2.4-fold, 2.92-fold to suffer more sever PE respectively. As a previous study revealed, patients with coronary heart disease have a higher prevalence of PE than those without cardiovascular disease which indicated similar risk factors of coronary heart disease and PE [17].
In addition, patients with coronary heart disease usually have lower cardiac reserve function. When occurred with PE, hemodynamics instability and elevated biomarkers such as TNI and NT-proBNP may increase the PE severity. As another type of VTE, DVT usually coexist with PE. The diagnosis of concomitant DVT has been identi ed as an independently risk factor of death within the rst 3 months after acute PE which indicated a more sever PE [18]. The underlying mechanism of respiratory failure caused by PE is the sever mismatch of ventilation and perfusion which results in a decreased diffusing capacity of the lungs. Furthermore, respiratory failure contributes to myocardial hypoxia injury and increase the expression of cardiac troponin which would increase the PE severity. Unfortunately, we didn't found association between comorbidities and cytokines levels (data not shown), which indicated that comorbidities do not affect PE severity through cytokine levels.
There are several limitations to our study that must be addressed. First, the small number of high-risk PE patients must be emphasized, bearing the risk of sample size error. Second, the retrospective design of the study limited the richness of the research. We didn't observe the dynamic changes of cytokines in the course of disease. A prospective, ideally designed study with physicians randomly blinded to routinely collected data would be required to solve the above problems.

Conclusion
IL-8 is negative correlated with PE severity. Patients with comorbidities regarding coronary heart disease, deep venous thrombosis, and respiratory failure may have higher risk to surfer a more sever PE.

Declarations Ethics approval and consent to participate
This study complied with the Declaration of Helsinki. The study protocol was approved by the Ethics Committee of Xinhua Hospital A liated to Shanghai Jiao Tong University School of Medicine (NO. XHEC-D-2021-170).

Consent for publication
Not applicable.

Availability of data and materials
The datasets used and/or analysed during the current study are available from Xiaoming Li (email: lixiaoming13734@126.com ) on reasonable request.

Competing interests
The authors declared no competing interests.

Funding
No funding.
Authors' contributions XML conceived the study and participated in its design and performance, statistical analysis, and drafting and revising the manuscript. YKL conceived the study and participated in its design and performance, statistical analysis, and drafting and revising the manuscript. WJY and XC conceived the study and participated in its design and performance. XJG conceived the study, participated in its design and coordination, and revised the manuscript. WG conceived the study, participated in its design and coordination, and revised the manuscript. All authors reviewed the manuscript.