|Year : 2022 | Volume
| Issue : 3 | Page : 239-244
Cytomorphological analysis of urinary cells among active and passive cigarette smokers in South-West, Nigeria
Ayodeji Blessing Ajileye1, Frederick Olusegun Akinbo2
1 Department of Biomedical Laboratory Science, College of Medicine, University of Ibadan, Ibadan, Nigeria
2 Depatment of Medical Laboratory Science, School of Basic Medical Sciences, University of Benin, Benin, Nigeria
|Date of Submission||10-Sep-2021|
|Date of Acceptance||21-Dec-2021|
|Date of Web Publication||15-Sep-2022|
Dr. Ayodeji Blessing Ajileye
Department of Biomedical Laboratory Science, College of Medicine, University of Ibadan, Ibadan
Source of Support: None, Conflict of Interest: None
Introduction: Cigarette smoke contains thousands of identified chemicals, among which sixty-nine (69) of these chemicals are known to be poisonous to humans and are capable of causing cancer in any part of the body. This study was conducted to evaluate the cytology of urinary cells among active and passive cigarette smokers in South-Western Nigeria. Materials and Methods: Urine samples were collected from 250 active cigarette smokers and 200 passive cigarette smokers who live in South-Western, Nigeria. The urine specimen was collected from participants and processed using the standard technique for microscopic examination. Smears were stained with haematoxylin and eosin and Papanicolaou stain. Results: The urinary smears of passive cigarette smokers revealed normal superficial squamous cells, transitional cells, cast, necrotic debris and mild inflammatory cells, while that of active cigarette smokers revealed high cellular turnover, protein casts, dysplastic epithelial cells and atypical urothelial cells with degenerative features. Conclusion: On the basis of this study, the cytomorphological analysis of urinary smears among active cigarette smokers revealed cytopathological features which connote the harmful effects of the chemical constituents present in cigarette smoke.
Keywords: Cigarette, cytocentrifuge, papanicolaou, squamous, urothelial
|How to cite this article:|
Ajileye AB, Akinbo FO. Cytomorphological analysis of urinary cells among active and passive cigarette smokers in South-West, Nigeria. Adv Hum Biol 2022;12:239-44
|How to cite this URL:|
Ajileye AB, Akinbo FO. Cytomorphological analysis of urinary cells among active and passive cigarette smokers in South-West, Nigeria. Adv Hum Biol [serial online] 2022 [cited 2022 Dec 2];12:239-44. Available from: https://www.aihbonline.com/text.asp?2022/12/3/239/356103
| Introduction|| |
Cigarettes are products consumed through smoking and manufactured out of cured and finely cut tobacco leaves. It is often combined with other additives, which are then rolled or stuffed into a paper-wrapped cylinder. Cigarettes are ignited and inhaled, usually through a cellulose acetate filter, into the mouth and lungs. Diseases that are related to cigarette smoking have claimed and are still claiming a large number of lives worldwide., Cigarette smoke affects the respiratory system, the circulatory system, the reproductive system, the skin, the eyes, and it increases the risk of many different cancers. This also includes those that are affected secondarily, such as babies born prematurely due to prenatal maternal smoking and victims of second-hand exposure to tobacco carcinogens. Second-hand smoke (passive smoke) is a mixture of smoke from the burning end of a cigarette and the smoke exhaled from the lungs of smokers. Second-hand smoke is involuntarily inhaled, lingers in the air for many hours after cigarettes must have been lightened, and this can cause a very wide range of adverse health effects, which include cancers, respiratory infections and asthma. Passive smokers who are exposed to second-hand smoke at work or at home increase their heart risk by 25%–30% and their lung cancer risk by 20%–30%. Cigarette smoke is a complex mixture of over 7000 identified chemicals; at least 250 of these identified chemicals are known to have specific toxicological properties, among which 69 are known to cause cancer and are also poisonous. The most important identified chemicals causing cancer are those that produce DNA damage since such damage appears to be the primary underlying cause of cancer. These cancer-causing chemicals include acrolein, formaldehyde, acrylonitrile, 1,3-butadiene, acetaldehyde, isoprene, aromatic amines, arsenic, benzene, benzo (α) pyrene, beryllium, cadmium, chromium, ethylene oxide, nickel polonium-210, polycyclic aromatic hydrocarbons and tobacco-specific nitrosamines.
The main addictive compound in cigarette smoke is nicotine., Nicotine belongs to the alkaloid family of compounds. Cigarette and other tobacco products vary widely in their content of nicotine, cancer-causing substances and other toxicants. The way an individual smokes a tobacco product is important as the nicotine content of the product determining how much nicotine gets into the body. Nicotine is absorbed into the bloodstream through the lining of the mouth and through the alveoli in the lungs and travels to the brain in a matter of seconds. Taking more frequent and deeper puffs of tobacco smoke increases the amount of nicotine absorbed by the body. Cigarette smoking has been seen to be the major risk factor for developing lung cancer, leukaemia, and cancer of the oesophagus, larynx, mouth, throat, liver, pancreas, stomach, cervix, colon, rectum, kidney and bladder. In addition, cigarette smoking causes inflammation and impairs immune function. The identified carcinogenic substances present in cigarette smoke escape the filtration process, and they become concentrated in the renal system. The effects of these chemicals are felt on the lining of the kidney down to the urinary bladder, alternately increasing the chances of developing bladder cancer. The risk of developing this cancer increases based on the number of cigarettes sticks an individual smokes in a day and the number of years an individual has been smoking. Several studies have laid much emphasis on the effects of cigarette smoke on the lungs, cervix, bladder, colon, stomach and pancreas. Information is lacking on the cytological analysis of urinary smears of cigarette smokers in South-Western Nigeria. The aim of this study was to evaluate the cytomorphology of urinary cells among active and passive cigarette smokers in South-Western Nigeria.
| Materials and Methods|| |
This study was conducted in the South-western states of Nigeria, namely Ondo, Ekiti, Osun, Oyo, Ogun and Lagos State. South-West is considered to be the most educationally advanced geopolitical zone in Nigeria and majorly a Yoruba-speaking region but with different Yoruba dialects among the people of the region. All the states in South-western Nigeria have common weather conditions of the dry season, which spans between October to February and raining season that is between March to October.
This study was carried out on 250 participants that only smoked a cigarette (active smokers) for at least 5 years and 200 subjects that did not smoke (passive smokers). Participants for this study were recruited from all the six South-Western states in Nigeria. The smokers were enrolled at various parks in the six states of South-Western Nigeria. The controls were apparently healthy, non-smoking that were recruited from schools and offices. The age range of participants ranged from 25 to 65 years and above. Participants that consented to participate and those males and females that have been smoking for 5 years and above were included in this study.
Participants that refused consent and those that are <5 years in smoking were excluded from this study. A structured questionnaire bothering on bio-data and socio-demographic characteristics was administered to participants prior to specimen collection. Informed consent was sought from each participant. The protocol for this study was approved by the ethics and research committees of Ministry of Health, Ado Ekiti; Osogbo, Osun State; Ibadan, Oyo State and Federal Medical Centre, Owo, Ondo State with the approval no MOH/EKHREC/EA/P/11; OSHREC/PRS/569T/181; AD13/479/4046B and OW/380/VOL. CX/150, respectively.
Sample size determination
Data from World Health Organization showed the incidence rate of cigarette smokers in men and women in Nigeria to be 18.1%. Therefore 18.1% prevalence was used to determine the sample size.
P (reported prevalence of cigarette smokers in Nigeria) = 0.181
The sample size for this research work will be determined using:
n = Required sample size
Z = Confidence level at 95% (standard value of 1.96)
P = Estimated prevalence (18.1%)
d = Accepted error
n = 228 (minimum sample size)
A total number of 450 participants that consisted of 250 active cigarette smokers and 200 passive cigarette smokers were recruited for this study.
Specimen collection and processing
About 20 ml of freshly voided urine specimen was collected from participants at the early hour of the day into sterile universal bottles and transported to the laboratory for processing. Urine smears (4) were made from the urine deposit following centrifugation and stained with haematoxylin and eosin and Papanicolaou staining techniques.
Two of the four smears made and fixed immediately were stained in haematoxylin and 0.5% Eosin solutions. Briefly, hydrated smears were stained in Harris haematoxylin for 4 min, rinsed in tap water and differentiated in 0.5% acid alcohol briefly. Smears were rinsed in water and blued in tap water for 10 min. Stained smears were counterstained with 0.5% eosin solution for 2 min, rinsed in water and dehydrated in ascending grades of alcohol, cleared in xylene and mounted with DPX.
The last two fixed smears were stained in the Papanicolaou staining technique. Hydrated smears were stained in Harris haematoxylin for 4 min, rinsed in tap water, differentiated in 0.5% acid alcohol for few seconds, rinsed in water and blued in tap water for 10 min. Stained smears were rinsed in 70% alcohol, 90% alcohol for 10 s and stained in orange G6 for 2 min. Smears were rinsed in 2 changes of 95% alcohol, stained in Eosin Azure 50 for 4 min and rinsed in 2 changes of 95% alcohol. Stained smears were dehydrated in two changes of absolute alcohol, cleared in xylene and mounted with DPX. The stained slides were examined using a ×40 objective lens.
Statistical Package for the Social Sciences (SPSS) software version 25 was the statistical package used in analyzing all data obtained in this study. Student t-test and Pearson Chi-square were used to comparing the means of the different analyses at P ≤ 0.05 statistical significance.
| Results|| |
Of the 450 subjects recruited, 385 (85.6%) were males, which consisted of 223 (89.2%) active male cigarette smokers, and 65 (14.4%) were females with 27 (10.8%) active female cigarette smokers. In addition, gender did not significantly affect the prevalence of active cigarette smokers (P = 0.171) [Table 1].
|Table 1: Distribution of active cigarette smokers according to their age groups|
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Out of 385 (85.6%) males recruited for this study, 162 (81%) were passive male smokers, while out of 65 (14.4%) females recruited for this study, 38 (19%) were passive female smokers. Furthermore, gender did not associate strongly with the prevalence of passive smokers in this study (P = 0.142) [Table 2].
|Table 2: Distribution of passive cigarette smokers according to their sex|
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Among the 250 active cigarette smokers recruited for this study, participants within the age group 25–34 years recorded the highest prevalence of 39.2%; followed by 35–44 years age group (37.6%); 45–54 years age group (11.6%); 55–64 years age group (6.8%); while subjects within the age group 65 years and above recorded the least prevalence of 4.8%. Among the 200 passive cigarette smokers, participants within the age group 25–34 years recorded the highest prevalence of 62%; followed by 35–44 years age group (25%); 45–54 years age group (7%); 55–64 and 65 years and above groups had 3% each. The age group significantly affected the prevalence of active and passive cigarette smokers in this study (P < 0.001) [Table 3].
|Table 3: Distribution of active and passive smokers according to their age groups|
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Cytomorphological analysis of urinary cells among active cigarette smokers revealed different cell types ranging from inflammatory cells with a prevalence of 93.6%, atypical cells (66.8%), cast (42%) and red blood cells (7.6%). At the same time, cytomorphological analysis of urinary cells of passive cigarette smokers revealed inflammatory cells with a prevalence of 24%, cast (17.6%), atypical cells (0.4%). No red blood cell was seen in the urinary cells of passive cigarette smokers. Being an active cigarette smoker influenced the production of inflammatory cells and atypia cells (P < 0.0001) [Table 4] significantly.
|Table 4: Morphological diagnosis of urinary smears of active and passive cigarette smokers|
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Among subjects who smoked ≤10 sticks of cigarette daily, 134 (23.6%) revealed infiltration of inflammatory cells; followed by 11–20 sticks of cigarette daily 83 (33.2%); while ≥21 sticks of cigarette daily, 17 (6.8%). The number of cigarette sticks smoked daily strongly influenced the presentation of infiltrated inflammatory cells in the urinary smears of active cigarette smokers (P < 0.05) [Table 5].
|Table 5: Relationship between numbers of cigarette sticks smoked daily and infiltration of inflammatory cells among active cigarette smokers|
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Among participants who smoked ≤ 10 sticks of cigarette daily, 73 (29.2%) revealed atypical urothelial cells, followed by 11–20 sticks of cigarette daily 77 (30.4%); while ≥21 sticks of cigarette daily, 17 (6.8%). The number of cigarette sticks smoked daily strongly influenced the presence of atypical urothelial cells in the urinary smears of active cigarette smokers (P < 0.0001) [Table 6].
|Table 6: Relationship between numbers of cigarette sticks smoked daily and atypical urothelial cells among active cigarette smokers|
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| Discussion|| |
Cigarette smoke contains thousands of identified chemicals, among which sixty-nine (69) of these chemicals are known to be poisonous to humans and are capable of causing cancer in any part of the body. The effects of cigarette smoke on the renal system of an active cigarette smoker depend greatly on the number of years an individual has been smoking and the number of cigarette sticks an individual can smoke per day. The kind of vicinity an individual lives in can determine the amount of second-hand smoke a passive cigarette smoker can inhale and accumulate over time.
Among the 250 active cigarette smokers recruited for this study, subjects within the age group of 25–34 years recorded the highest prevalence (39.2%) of smokers; followed by 35–44 years (37.6%), 45–54 years (11.6%), 55–64 years (6.8%) and the least prevalence (4.8%) was recorded among 65 years and above. This finding is in agreement with Action on Smoking and Health, which observed a high prevalence of smoking among young adults that are within the age group of 20–35 years and continues to be lowest among the age group 65 years and above. This study observed a strong significant difference between the prevalence of active cigarette smokers when compared with that of passive cigarette smokers (P < 0.0001). This trend shows that the majority of the subjects started smoking at a younger age and became addicted to it, which may be partly caused by peer group influence.
The number of cigarette sticks smoked daily among active cigarette smokers varies widely, with 148 (59.2%) of the subjects being the highest, smoked ≤10 sticks of cigarette daily; followed by 85 (34%) smoked 11–20 sticks of cigarette daily; while 17 (6.8%) smoked ≥21 sticks of cigarette daily being the lowest. This finding is in agreement with a study conducted by Inyang et al. that observed that subjects who smoked ≤10 sticks of cigarette daily were 47.2% and subjects who smoked 11–15 sticks of cigarette daily were 27.1%. According to Cooper, the adverse effects of cigarette smoke on the body system depends on the number of years an individual may have been smoking and the number of cigarette stick that is consumed in a day.
Smoking cigarettes early in life or being exposed to second-hand cigarette smoke early in life and for a long time can increase the risk of an individual developing cancer in the renal system or any organ of the body. The urine cytology of passive cigarette smokers [Figure 1] revealed superficial squamous cells, intermediate squamous cells, transitional cells, cast and necrotic debris. Mild infiltrate of inflammatory cells were also observed in some of the urinary smears of passive cigarette smokers. This may be due to the fact that these individuals must have stayed, worked or lived in the vicinity where people smoke cigarettes. The environment a passive cigarette smoker lives or works in, the kind of peer groups, and the number of years of exposure to second-hand cigarette smoke has a role in determining the hazardous effects of cigarette smoke on the renal system of such individual. Nigeria as a country has very weak regulations and enforcement on the sales, promotion and advertisement of cigarettes, as it is sold in open places both in rural and urban centres. There is a law regulating where an individual can smoke in Nigeria, but this law is not fully enforced. Smokers puff up cigarette sticks in open places and where people are gathered and not minding the effects of this second-hand smoke on people who are non-smokers.
|Figure 1: Photomicrographs: Urine smear of a passive cigarette smoker showing (A) normal superficial squamous cells (H and E, ×100).|
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About 70% of the urinary smears [Figure 2] & [Figure 3] from active cigarette smokers revealed heavy infiltration of inflammatory cells. This finding is consistent with the previous studies of Ajileye et al., who observed the same among 70% of cigarette smokers in Owo town, Ondo State and Inyang et al. in Calabar metropolis. This finding may be due to the fact that the toxic chemicals present in the cigarette smoke escape the filtration process in the kidneys due to the damages caused by the toxic chemicals, pass through the ureter and settle in the urinary bladder, thereby damaging the cells and epithelial lining the ureter, urinary bladder and the urethra.
|Figure 2: Photomicrographs: Urine smear from an active cigarette smoker, who smokes 11–20 cigarette sticks daily, showing (A) atypical urothelial cells with an increased nucleo-cytoplasmic ration (B) heavy infiltration of inflammatory cells (C) red blood cell (H and E, ×100).|
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|Figure 3: Photomicrographs: Urine smear from an active cigarette smoker who smokes ≥21 cigarette sticks daily, showing (A) Atypical urothelial cells with enlarged nucleus (B) red blood cell (C) infiltrates of inflammatory cells (H and E, ×100).|
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The urine smears of active cigarette smokers [Figure 4], [Figure 5], [Figure 6] revealed atypical urothelial cells, degenerative cells, red blood cells, pleomorphisms, hyperchromatic cells and infiltration of inflammatory cells. This is consistent with the previous study of Azhar et al. that was reported in other urothelial cells seen in the smears of active cigarette smokers include protein cast, moderate red blood cells, atypical urothelial cells, degenerative cells, increased in nucleo-cytoplasmic ratio and coarse chromatin. This finding is in agreement with Inyang et al. The various degrees of cytomorphological features observed in the smears of active cigarette smokers may be due to the fact that the heavy metals, tars, formaldehyde and other carcinogenic substances present in cigarette smoke are capable of affecting the epithelial lining of the renal system, thereby altering the normal morphology of the urinary cells which can eventually lead to renal cell lesions based on the number of sticks of cigarette is taken and duration of smoking.
|Figure 4: Photomicrographs: Urine smear from an active cigarette smoker who smokes ≤10 cigarette sticks daily, revealing (A) transitional cell with an increased nucleo-cytoplasmic ratio (B) Inflammatory cells (H and E, ×40).|
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|Figure 5: Photomicrographs: Urine smear from an active cigarette smoker who smokes 11–20 cigarette sticks daily, showing (A) degenerating cells with increased nucleo-cytoplasmic ratio, hyperchromatic and suspicious for malignancy (H and E, ×100).|
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|Figure 6: Photomicrographs: Urine smear from an active cigarette smoker, who smokes ≥20 cigarette, sticks daily, showing (A) inflammatory cells (B) Atypical urothelial cells with features of increased nucleo-cytoplasmic ratio and hyperchromatic (H and E, ×100).|
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Active cigarette smokers [Figure 2] who usually smoke ≤10 sticks of cigarette daily for a minimum of 5 years did not reveal many cytopathological features in their urinary cells compared to those who usually smoke ≥11 sticks of cigarette daily. This can be attributed to the fact that the higher the number of sticks of cigarettes consumed a day, the greater the degree of toxicity on the renal system. This is a strong indication that the carcinogenic substances present in cigarette smoke have seriously distorted the cell morphology and thereby caused serious injury to the renal system of these active cigarette smokers. The degree of this injury is based on the number of cigarette sticks an individual smokes per day and the number of years an individual has been engaged in cigarette smoking.
These abnormal features are caused as a result of the cytotoxic effects of the harmful chemicals and carcinogenic substances that are present in cigarette smoke. Governments and Non-Governmental Organizations should mount health education programs on social media on the implication of what cigarette smoke can cause to the entire organs of the body and most especially the renal system.
| Conclusion|| |
The prevalence of male active cigarette smokers (89.2%) was higher than that of female active cigarette smokers (10.8%). There is a high prevalence (39.2%) of cigarette smoking among youths in South-Western Nigeria. Cytomorphological analysis of urinary cells among active cigarette smokers revealed different cell types ranging from infiltrating of inflammatory cells, atypical cells, cast and red blood cells. These abnormal features are caused as a result of the cytotoxic effects of the harmful chemicals and carcinogenic substances that are present in cigarette smoke.
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Conflicts of interest
There are no conflicts of interest.
| References|| |
Warren GW, Kasza KA, Reid ME, Cummings KM, Marshall JR. Smoking at diagnosis and survival in cancer patients. Int J Cancer 2013;132:401-10.
Hajek P, Etter JF, Benowitz N, Eissenberg T, McRobbie H. Electronic cigarettes: Review of use, content, safety, effects on smokers and potential for harm and benefit. Addiction 2014;109:1801-10.
Arcavi L, Benowitz NL. Cigarette smoking and infection. Arch Intern Med 2004;164:2206-16.
Wright ME, Michaud DS, Pietinen P, Taylor PR, Virtamo J, Albanes D. Estimated urine pH and bladder cancer risk in a cohort of male smokers (Finland). Cancer Causes Control 2005;16:1117-23.
Travis LB, Rabkin CS, Brown LM, Allan JM, Alter BP, Ambrosone CB, et al.
Cancer survivorship – Genetic susceptibility and second primary cancers: Research strategies and recommendations. J Natl Cancer Inst 2006;98:15-25.
Djordjevic MV, Doran KA. Nicotine content and delivery across tobacco products. Handb Exp Pharmacol 2009;61-82.
Henningfield JE, Fant RV, Radzius A, Frost S. Nicotine concentration, smoke pH and whole tobacco aqueous pH of some cigar brands and types popular in the United States. Nicotine Tob Res 1999;1:163-8.
Walter V, Jansen L, Hoffmeister M, Brenner H. Smoking and survival of colorectal cancer patients: Systematic review and meta-analysis. Ann Oncol 2014;25:1517-25.
Peto R, Darby S, Deo H, Silcocks P, Whitley E, Doll R. Smoking, smoking cessation, and lung cancer in the UK since 1950: Combination of national statistics with two case-control studies. BMJ 2000;321:323-9.
Hodges KB, Lopez-Beltran A, Emerson RE, Montironi R, Cheng L. Clinical utility of immunohistochemistry in the diagnoses of urinary bladder neoplasia. Appl Immunohistochem Mol Morphol 2010;18:401-10.
Naing L, Winn T, Rusli BN. Practical issues in calculating sample size for prevalence studies. Arch Orofac Sci 2006;2:9-14.
Ochei J, Kolhatkar A. Medical Laboratory Science Theory and Practice. Tata McGraw-Hill Publishing Company Limited, New Delhi; 2005. p. 450-521.
Jhee JH, Joo YS, Kee YK, Jung SY, Park S, Yoon CY, et al.
Secondhand smoke and CKD. Clin J Am Soc Nephrol 2019;14:515-22.
Action on Smoking and Health. Smoking Statistics; Who Smokes and How Much; 2016. Available from: https://www.ash.org.uk
. [Last accessed on 2018 Mar 15].
Inyang IJ, Bassey IE, Udonkang M, Ugori C, Udoka C. Urine cytology screening for bladder cancer among tobacco cigarette smokers in Calabar metropolis. Int J Med Lab Res 2018;3:52-8.
Cooper RG. Effects of tobacco smoking on renal function. Indian J Med Res 2006;124:261-68.
] [Full text]
Ajileye AB, Eze GI, Fasogbon SA. Cytology analysis of urine among cigarette smokers. Am J Biomed Sci 2016;8:56-67.
Azhar H, Parmvir D, Megan N. R, Carla A, Vishal S. Neoplastic Pathogenesis Associated with Cigarette Carcinogens. Cureus 2019; 11(1): e3955.
Khalil MA, Tan J, Khamis S, Khalil MA, Azmat R, Ullah AR. Cigarette smoking and its hazards in kidney transplantation. Adv Med 2017;2017:6213814.
Chen H, Xu C, Jin Q, Liu Z. S100 protein family in human cancer. Am J Cancer Res 2014;4:89-115.
[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6]
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6]