ABSTRACT

Objective:

Smoking causes endothelial dysfunction by causing systemic inflammation. Our aim is to investigate the predictive role of fibrinogen albumin ratio (FAR), neutrophil lymphocyte ratio, platelet lymphocyte ratio (PLR), monocyte lymphocyte ratio (MLR), monocyte high-density lipoprotein ratio (MHR), mean platelet volume (MPV), systemic immune inflammatory index (SII) and systemic inflammatory response index (SIRI) which are the newest markers of systemic inflammation, in smoking.

Method:

The study was planned as a single-center, prospective case-control study. People aged between 20-45 years [n=76; female/male (n)=37/39] who met the inclusion criteria were included. Two groups consisting of smokers (n=38) and non-smokers (n=38) were compared according to demographic data and biochemical values. p-value was accepted as <0.05 in terms of statistical significance.

Results:

The proportion of men in the study group was 51.3% [total n=76; mean age 30.18±5.75 years; female/male (n):37/39]. There was no significant difference between the two groups of smokers (n=38) and non-smokers (n=38) in terms of gender and age (p=0.386, p=0.296, respectively). The average amount of smoking was 22.84±11.48 packs/year and 29.73±9.94 packs/day. FAR, MPV, MHR, MLR and SIRI were significantly higher and PLR was significantly lower in the smokers than the other group (p<0.05). According to the ROC analysis, the areas under the curve of FAR, MHR, MLR, SIRI, MPV and PLR were determined as 0.70, 0.90, 0.83, 0.80, 0.69 and 0.66, respectively (p<0.001). The variants with the highest sensitivity were monocytes and MPV, and the variant with the highest specificity was monocytes. FAR was positively correlated with MHR, MLR, SIRI (p=0.00, r=0.34; p=0.05, r=0.2; p=0.03, r=0.24, respectively).

Conclusion:

Our study suggests that new inflammatory markers can be used to predict the inflammatory process in endothelial dysfunction caused by smoking. It may be beneficial to conduct new studies with larger number of participants to see whether these markers can also be predictors of other silent inflammatory processes in non-smokers.

Keywords: Endothelial dysfunction, fibrinogen albumin ratio, inflammation, monocyte, smoking

Introduction

Endothelial dysfunction is defined as decreased nitric oxide synthesis and impairment of relaxation-contraction functions (1). Cardiovascular risk factors such as smoking, diabetes, atherogenic dyslipidemia, abdominal obesity, hypertension and age initiate the inflammatory process by disrupting the endothelial structure (2,3). Atherosclerosis is a chronic and progressive disease characterized by endothelial dysfunction and inflammation. Deterioration of vascular tone, increased cytokine production, accumulation of inflammatory cells in the endothelium, and migration of smooth muscle cells to the damaged area cause atherosclerotic plaque formation (4).

Smoking causes disorders in the hemostatic system and lipid profile and endothelial dysfunction (5). It also predisposes the body to thrombosis and disrupts the procoagulant fibrinolytic activity of the endothelium (6). While smoking suppresses anti-inflammatory cytokines, it causes systemic inflammation in the body by using pro-inflammatory cytokines and acute phase proteins such as C-reactive protein (CRP) and fibrinogen (7).

The increase in plasma viscosity is associated with cardiovascular diseases because it leads to a procoagulant state. The main substances that increase viscosity are fibrinogen and lipoproteins. Fibrinogen may play a triggering role in cardiovascular diseases by increasing viscosity or by affecting the platelets and fibrin formation mechanism. Increased fibrinogen in chronic smokers contributes to the inflammatory and atherosclerotic basis (8). In a prospective study, it was shown that increased fibrinogen and CRP have an effect on the pathogenesis of cardiovascular diseases (9,10).

Normal albumin levels have antiatherogenic and anti-inflammatory effects in the body. Therefore, the decrease in albumin causes an increase in pro-inflammatory markers along with an increase in systemic inflammation in the body and is linked to oxidative stress (11). In a study, it was shown that serum albumin value varies in smokers and has an important role in the development of coronary artery disease in those people (12). In subsequent studies, serum albumin was shown to be a strong risk factor for MI when other risk factors were adjusted (13).

Studies have reported that high fibrinogen/albumin ratio (FAR) is an important marker for inflammation and atherosclerosis (9). Neutrophil/lymphocyte ratio (NLR) levels have been found to be quite high in diseases that cause endothelial damage, such as diabetes, myocardial infarction, atherosclerosis and hypertension, and are considered a poor prognostic factor (14). The diagnostic role of monocyte/lymphocyte ratio (MLR) in showing the level of inflammation in cardiovascular and autoimmune diseases has been proven (15). Monocyte/high-density lipoprotein (HDL) ratio (MHR) is emerging as a new prognostic indicator in predicting oxidative stress and systemic inflammation levels in the atherosclerosis process (16). Platelet/lymphocyte ratio (PLR), which is an important indicator of inflammation and thrombosis, is used in many diseases (17). Mean platelet volume (MPV) indicates platelet activation in vascular diseases (18). The role of inflammation in all stages of cancer has been proven, and the increase in systemic inflammatory response index (SIRI) levels, an important inflammatory marker, has been shown to be valuable in predicting survival in cancer patients (19). Systemic immune inflammatory index (SII), a new marker, shows the immune response and inflammation in the body more strongly than other markers (20).

In this study, we aimed to investigate FAR, a new inflammatory marker, as an indicator of endothelial dysfunction due to smoking. In addition, it is planned to study MLR, NLR, PLR, MPV, MHR, SII and SIRI as inflammatory markers in smokers.

Materials and Methods

Study group

This study was planned as a single-center, prospective case-control study. Seventy-six participants aged 20-45 years, who applied to the outpatient clinics between November 2021 and April 2022, were included according to inclusion criteria. Individuals who had chronic lung, liver and kidney diseases, connective tissue diseases, acute or chronic infection, malignancy, metabolic syndrome, obesity, pregnancy status, menopause, chronic diseases such as hypertension and diabetes mellitus, severe trauma or surgery in the last 6 months, cardiac disease, any chronic medication, Coronavirus disease-2019 in the last 6 months and active alcohol users were excluded. Demographic data and biochemical values of the groups were compared among two groups as smokers (n=38) and non-smokers (n=38).

Ethics Committee Approval

This study was approved (dated: 06/03/2020 and numbered: 2213) by Clinical Research Ethics Committee of University of Health Sciences Turkey, İstanbul Training and Research Hospital. The expenses of the study were covered by University of Health Sciences Turkey, İstanbul Training and Research Hospital. All participants were informed about the study and their consent was obtained.

Sample Size

Cohen’s d effect size was 0.80; α value 0.05; the power (1-β) value was taken as 0.80, and the minimum number of samples required for statistical comparison of two independent groups was calculated as n=30 participants for each group through the G*Power version 3.1.9.4 (Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany).

Statistical Analysis

Statistical analyzes were performed with the help of SPSS version 28.0 program. To assess the normality of the data, Kolmogorov-Smirnov test was used. Mean, standard deviation and median values were used when presenting descriptive analyzes. Two-sample independent test was used for parametric variables and Mann-Whitney U test was used for non-parametric variables. Receiver operating characteristic (ROC) analysis was applied to evaluate the performance of the tests with diagnostic accuracy, sensitivity, specificity and area under the curve (AUC) values. Pearson Correlation test was used to evaluate the relationships between quantitative variables by accepting the variables that had a significant relationship with the FAR as independent variables. p-value was accepted as <0.05 in terms of statistical significance.

Results

Participants consisted of 37 women and 39 men. Two groups were formed as smokers (n=38) and non-smokers (n=38). The mean age was 29.78±5.75 years. The average height was 170.22±8.51 cm, the average weight was 69.44±12.62 kg and body mass index (BMI) were 23.86 kg/m². No statistically significant difference was found between the demographic data of the two groups (p>0.05) (Table 1). The smoking rates of the individuals were 22.84±11.48 pack/year and 29.73±9.94 pcs/day. No statistically significant difference was observed when smoking amounts of smokers were compared according to gender (p>0.05) (Table 2).

Table 1

Table 2

The levels of white blood cell (WBC), neutrophil, monocytes, lymphocyte, MPV, fibrinogen, MHR, MLR, SIRI and FAR were significantly higher in smokers compared to non-smokers (p<0.05). Albumin and PLR levels were found to be significantly lower in the smokers (p<0.05). HDL values were found to be significantly higher in non-smokers, while triglyceride (TG) and low-density lipoprotein (LDL) values were significantly higher in smokers (p<0.05) (Table 3).

Table 3

According to the ROC analysis, the AUC of FAR, MHR, MLR, SIRI, MPV and PLR were determined as 0.70, 0.90, 0.83, 0.80, 0.69 and 0.66, respectively (p<0.001) (Table 4). Monocyte had the highest sensitivity (90.48%) and specificity (92.30%) among all of the inflammatory markers when cut-off point was calculated as 0.50 10⁹/L. The AUC for monocytes was 93.7% (95% confidence interval: 0.60-0.83) (p<0.001) (Table 4, Figure 1).

Table 4

Figure 1

A significant positive correlation was found between FAR and the new inflammatory markers MHR, MLR and SIRI (p=0.00, r=0.34; p=0.05, r=0.22; p=0.03, r=0.24) (Table 5).

Table 5

Discussion

It is known that fibrinogen is one of the most important factors determining plasma viscosity. As fibrinogen levels increases, flow of blood cells slows down, they tend to stick together and viscosity increases. These events cause atherosclerotic changes in vessels. As a result, it has been shown that the increase in plasma fibrinogen levels is effective in the process starting with endothelial dysfunction and progressing to atherosclerosis (21-23). Albumin is decreased in inflammation and is associated with high mortality in cardiovascular diseases (24). In the British Regional Heart Study, it was found that low albumin levels were associated with smoking (25). Karahan et al. (26) found FAR levels more sensitive compared to fibrinogen or albumin values alone in predicting venous insufficiency. In another study, FAR was found to be high in patients with stable angina pectoris, which was associated with lower mortality (27).

In our study, in accordance with the literature, fibrinogen levels were found to be high and albumin levels were low in smokers (25). FAR, one of the markers we used to predict endothelial dysfunction in smoking, was found to be high in the smoking group. The early stage of atherosclerosis is endothelial dysfunction, and previous studies have shown that FAR is high in atherosclerosis. In conclusion, our study suggests that high FAR may indicate endothelial dysfunction.

In a study, WBC, neutrophil and lymphocyte counts were found to be high in smokers (28). Similar effects of smoking were revealed in the study of Gumus et al. (29). In the study of Kutlu and Demirbas (30) MPV was found to be significantly higher in smoking individuals while no significant difference was found in platelet count. In our study, WBC, neutrophil, lymphocyte, monocytes and MPV levels were found to be significantly higher in the smokers group. It is well known that platelets are involved in the atherosclerotic process and MPV shows the platelet activation. In our study, platelet count not statistically significant difference between groups. But MPV levels were found to be significantly higher in smokers, similar to the studies that examining the hemogram parameters of smokers (18). When we evaluated the parameters that increase with smoking, MPV was found to be one of the parameters with the highest sensitivity. Our study suggests that high MPV levels predict endothelial dysfunction in smokers.

NLR is a biomarker that has gained importance in showing morbidity and mortality in the atherosclerotic process (14). In the study by Çekici et al. (31), when two groups consisting of smokers and non-smokers were compared, the NLR value was found to be higher in the smokers group. In our study, NLR levels were found to be higher in smokers compared to the other group, but this elevation was not statistically significant. As reported above, our study group consisted of people without any comorbidities and lower mean age unlike other studies. Besides, few number of participants of our study may have caused this difference.

PLR is an indicator of inflammation in endothelial dysfunction and is also used as a marker for atherosclerosis and cardiovascular disease (17). In our study, PLR values of the two groups were compared and it was found to be significantly lower in the smoking group consistent with the study of Gumus et al. (29).

Demirbaş et al. (32), analyzed MHR, MLR and PLR as new inflammatory markers showing inflammation and oxidative stress in vitiligo and found them significantly higher in vitiligo patients than controls. Therefore these parameters were suggested as new indicators of inflammation in vitiligo patients (32). Many studies have investigated MHR in various diseases. MHR has gained importance in demonstrating systemic inflammation. High MHR is an increased risk for cardiovascular diseases (33). It has been shown that, in atherosclerosis the number of monocytes increase whereas HDL level remains low. We showed that the number of monocytes was significantly higher in the smoking group (p<0.05). According to ROC analysis, the sensitivity and specificity of monocyte was found to be the highest variable among the parameters increasing due to smoking. It is known that monocytes play a role in the first step of the atherosclerotic process and the number of monocytes increase in cardiovascular diseases (15,16). As the levels of MHR and MLR which are suggested as new inflammatory markers are evaluated, both were found to be significantly higher in the smokers group (p<0.05). The cut-off value we obtained in the ROC analysis was calculated as 0.09 for MHR and 0.20 for MLR. According to the results, we think that these two markers may be stimulatory and/or predictive in endothelial dysfunction due to smoking.

In a study, it was found that SIRI values of 285 patients with nasopharyngeal cancer were more significant in predicting the survival of patients compared to cancer staging (34). In another study, SIRI values were found to be an important predictor of survival in patients with pancreatic cancer (35). When other inflammation markers were examined, it was observed that they were not as strong as SIRI (36). This marker has been mostly investigated to determine the prognosis of cancer patients. The relationship between smoking and SIRI has not been investigated before, and our study shows the relationship between SIRI and smoking-induced inflammation.

Studies have shown that systemic inflammation has a very important role in the formation of cancer cells (37). SII has been found to be a strong predictor of survival for patients diagnosed with colorectal cancer. It was even found important in distinguishing high-risk among patients with the same cancer stage. In this study, SII was found to be more valuable when compared to other inflammation markers (38). In another study, many parameters were examined and NLR, PLR and SII were found to be significantly higher in patients with kerataconus. Here, the reason for the high SII was thought to be the role of endothelial dysfunction rather than chronic inflammation (39). In our study, SII was found to be high in smokers, but high SII was not statistically significant in demonstrating smoking-related endothelial dysfunction.

We think that this issue should be examined with more number of participants and comprehensive studies.

Conclusion

We showed that biomarkers such as FAR, MPV, PLR, MHR, MLR and SIRI may predict endothelial damage caused by smoking. Our study may inspire new and exhaustive ones investigating new inflammatory markers to predict silent inflammatory processes in healthy individuals.

Ethics

Ethics Committee Approval: This study was approved (dated: 06/03/2020 and numbered: 2213) by Clinical Research Ethics Committee of University of Health Sciences Turkey, İstanbul Training and Research Hospital.

Informed Consent: All participants were informed about the study and their consent was obtained.

Peer-review: Externally and internally peer-reviewed.

Authorship Contributions

Concept: Ş.A., H.F.A., Z.A.S., Design: Ş.A., H.F.A., Z.A.S., Data Collection or Processing: Ş.A., H.F.A., Analysis or Interpretation: Ş.A., H.F.A., Z.A.S., Drafting Manuscript: Ş.A., H.F.A., Critical Revision of Manuscript: Ş.A., H.F.A., Z.A.S., Final Approval and Accountability: Ş.A., H.F.A., Z.A.S., Supervision: Ş.A., H.F.A., Z.A.S., Writing: Ş.A., H.F.A., Z.A.S.

Conflict of Interest: No conflict of interest was declared by the authors.

Financial Disclosure: The expenses of the study were covered by University of Health Sciences Turkey, İstanbul Training and Research Hospital.

References

Bonetti PO, Lerman LO, Lerman A. Endothelial dysfunction: a marker of atherosclerotic risk. Arterioscler Thromb Vasc Biol 2003;23(2):168-175.

Behrendt D, Ganz P. Endothelial function. From vascular biology to clinical applications. Am J Cardiol 2002;90(10C):40L-48L.

Cornwell TL, Arnold E, Boerth, NJ, Lincoln TM. Inhibition of smooth muscle cell growth by nitric oxide and activation of cAMP-dependent protein kinase by cGMP. Am J Physiol 1994;267(5Pt1):C1405-C1413.

Sitia S, Tomasoni L, Atzeni F, Ambrosio G, Cordiano C, Catapano A, et al. From endothelial dysfunction to atherosclerosis. Autoimmun Rev 2010;9(12):830-834.

Esper RJ, Nordaby RA, Vilariño JO, Paragano A, Cacharrón JL, Machado RA. Endothelial dysfunction: a comprehensive appraisal. Cardiovasc Diabetol 2006;5:4.

Hung J, Lam JYT, Lacoste L, Letchacovski G. Cigarette smoking acutely increases platelet thrombus formation in patients with coronary artery disease taking aspirin. Circulation 1995;92:2432-2436.

Yanbaeva DG, Dentener MA, Creutzberg EC, Wesseling G, Wouters EF. Systemic effects of smoking. Chest 2007;131(5):1557-1566.

Alkan FA, Cakmak G, Karis D, Sağlam ZA, Saler T, Temiz LU, et al. The evaluation of plasma viscosity and endothelial dysfunction in smoking individuals. Clin Hemorheol Microcirc 2014;58(3):403-413.

Kayapinar O, Ozde C, Kaya A. Relationship Between the Reciprocal Change in Inflammation-Related Biomarkers (Fibrinogen-to-Albumin and hsCRP-to-Albumin Ratios) and the Presence and Severity of Coronary Slow Flow. Clin Appl Thromb Hemost 2019;25:1076029619835383.

Danesh J, Whincup P, Walker M, Lennon L, Thomson A, Appleby P, et al. Low grade inflammation and coronary heart disease: prospective study and updated meta-analyses. BMJ 2000;321(7255):199-204.

Don BR, Kaysen GA. Serum albumin: relationship to inflammation and nutrition. Semin Dial 2004;17(6):432-437.

Nelson JJ, Liao D, Sharrett AR, Folsom AR, Chambless LE, Shahar E, et al. Serum albumin level as a predictor of incident coronary heart disease: the Atherosclerosis Risk in Communities (ARIC) study. Am J Epidemiol 2000;151(5):468-477.

Yang Q, He YM, Cai DP, Yang XJ, Xu HF. Risk burdens of modifiable risk factors incorporating lipoprotein (a) and low serum albumin concentrations for first incident acute myocardial infarction. Sci Rep 2016;6:35463.

Sen N, Afsar B, Ozcan F, Buyukkaya E, Isleyen A, Akcay AB, et al. The neutrophil to lymphocyte ratio was associated with impaired myocardial perfusion and long term adverse outcome in patients with ST-elevated myocardial infarction undergoing primary coronary intervention. Atherosclerosis 2013;228(1):203-210.

Zuo B, Zhu S, Meng X, Zhao D, Zhang J. Monocyte/lymphocyte ratio is associated with carotid stenosis in ischemic stroke: A retrospective analysis. Brain Behav 2019;9(10):e01429.

Kundi H, Gok M, Kiziltunc E, Cetin M, Cicekcioglu H, Cetin ZG, et al. Relation Between Monocyte to High-Density Lipoprotein Cholesterol Ratio With Presence and Severity of Isolated Coronary Artery Ectasia. Am J Cardiol 2015;116(11):1685-1689.

Balta S, Ozturk C. The platelet-lymphocyte ratio: A simple, in expensive and rapid prognostic marker for cardiovascular events. Platelets 2015;26(7):680-681.

Tsiara S, Elisaf M, Jagroop IA, Mikhailidis DP. Platelets as predictors of vascular risk: is there a practical index of platelet activity? Clin Appl Thromb Hemost 2003;9(3):177-190.

Colotta F, Allavena P, Sica A, Garlanda C, Mantovani A. Cancer-related inflammation, the seventh hallmark of cancer: link stogenetic instability. Carcinogenesis 2009;30(7):1073-1081.

Hu B, Yang XR, Xu Y, Sun YF, Sun C, Guo W, et al. Systemic immune-inflammation index predicts prognosis of patients after curative resection for hepatocellular carcinoma. Clin Cancer Res 2014;20(23):6212-6222.

Van Breugel HF, de Groot PG, Heethaar RM, Sixma JJ. Role of plasma viscosity in platelet adhesion. Blood 1992;80(4):953-959.

Schmid-Schönbein H. Microrheology of erythrocytes, blood viscosity, and the distribution of blood flow in the microcirculation. Int Rev Physiol 1976;9:1-62.

Mannucci PM. Recent progress in the pathophysiology of fibrinogen. Eur Heart J 1995;16(Suppl A):25-30.

Asayama K, Uchida N, Nakane T, Hayashibe H, Dobashi K, Amemiya S, et al. Antioxidants in the serum of children with insulin-dependent diabetes mellitus Free Radic Biol Med 1993;15(6):597-602.

Falk E, Fuster V. Atherogenesis and its Determinants. In: Fuster V, Alexander RW, O’Rourke RA (editors). Hurst’s The Heart. 10th ed. USA. Division; 2001. Ch35, pp. 1065-1093.

Karahan O, Yavuz C, Kankilic N, Demirtas S, Tezcan O, Caliskan A, et al. Simple blood tests as predictive markers of disease severity and clinical condition in patients with venous insufficiency. Blood Coagul Fibrinolysis. 2016;27(6):684-690.

Dr. Leyla Çiftçi Tıpta Uzmanlık Tezi, Stabil Anjinalı Hastalarda Koroner Arter Hastalığı Ciddiyeti ile Fibrinojen/Albumin Oranı Arasındaki İlişkinin Araştırılması, Dicle, 2016

Luetragoon T, Rutqvist LE, Tangvarasittichai O, Andersson BÅ, Löfgren S, Usuwanthim K, et al. Interaction among smoking status, single nucleotide polymorphisms and markers of systemic inflammation in healthy individuals. Immunology 2018;154(1):98-103.

Gumus F, Solak I, Eryilmaz MA. The effects of smoking on neutrophil/lymphocyte, platelet/lymphocyte ratios. Bratisl Lek Listy 2018;119(2):116-119.

Kutlu R, Demirbas N. The Effects of Smoking on Platelet Count, Mean Platelet Volume and Cardiovascular Risk Factors: A Case-control Study. Med Bull Haseki 2017;55(4):299-305.

Çekici, Y, Yılmaz, M, Seçen, Ö. New inflammatory indicators: association of high eosinophil-to-lymphocyte ratio and low lymphocyte-to-monocyte ratio with smoking. J Int Med Res 2019;47(9):4292-4303.

Demirbaş A, Elmas ÖF, Atasoy M, Türsen Ü, Lotti T. Can monocyteto HDL cholesterol ratio and monocyte to lymphocyte ratio be markers for inflammation and oxidative stress in patients with vitiligo? A preliminary study. Arch Dermatol Res 2021;313(6):491-498.

Ganjali S, Gotto AM Jr, Ruscica M, Atkin SL, Butler AE, Banach M, et al. Monocyte-to-HDL-cholesterol ratio as a prognostic marker in cardiovascular diseases. J Cell Physiol 2018;233(12):9237-9246.

Chen Y, Jiang W, Xi D, Chen J, Xu G, Yin W, et al. Development and validation of nomogram based on SIRI for predicting the clinical outcome in patients with nasopharyngeal carcinomas. J Investig Med 2019;67(3):691-698.

Pacheco-Barcia V, Mondéjar Solís R, France T, Asselah J, Donnay O, Zogopoulos G, et al. A systemic inflammation response index (SIRI) correlates with survival and predicts oncological outcome form FOLFIRINOX therapy in metastatic pancreatic cancer. Pancreatology 2020;20(2):254-264.

Topkan E, Kucuk A, Ozdemir Y, Mertsoylu H, Besen AA, Sezen D, et al. Systemic Inflammation Response Index Predicts Survival Outcomes in Glioblastoma Multiforme Patients Treated with Standard Stupp Protocol. J Immunol Res 2020;2020:8628540.

Wang Y, Li Y, Chen P, Xu W, Wu Y, Che G. Prognostic value of the pretreatment systemic immune-inflammation index (SII) in patients with non-small cell lung cancer: a meta-analysis. Ann Transl Med 2019;7(18):433.

Chen JH, Zhai ET, Yuan YJ, Wu KM, Xu JB, Peng JJ, et al. Systemic immune-inflammation index for predicting prognosis of colorectal cancer. World J Gastroenterol 2017;23(34):6261-6272.

Elbeyli A, Kurtul BE. Systemic immune-inflammation index, neutrophil-to-lymphocyte ratio, andplatelet-to-lymphocyte ratio levels areas sociated with keratoconus. Indian J Ophthalmol 2021;69(7):1725-1729.

Plasma Fibrinogen to Albumin Ratio as an Indicator of Endothelial Dysfunction in Smoking