Waterpipe tobacco smoke toxicant exposure and effects

Alan ShihadehAmerican University of Beirut

1st International Conference on WTSOctober 20-23, 2013

Abu Dhabi

Waterpipe toxicants & health effects

http://firefighterparamedicstories.blogspot.com/2011_10_01_archive.html

What happens to the user?• acute physiological effects• BP, HR, inflammatory responses,

lung function…

Does waterpipe smoking emit toxicants?

exhaled mainstream (EMS)

sidestream (SS)

SS + EMS

Does the smoke contain toxicants?• particle size• chemistry• biological activity

Does the user absorb toxicants?biomarkers in: blood, breath, urine

absorbed

exhaledinhaled

What happened to the smoker?• long term mortality & morbidity

Molecule Particle Cell Animal Human individual Population health

hose

mouthpiece

body

bowl

head

coal

tobacco

water

Narghile waterpipe

Photo credit: internet, source unknown

hose

mouthpiece

body

bowl

head

coal

tobacco

water

Narghile waterpipe

Differences with cigarette

tobacco burn not self-sustaining charcoal needed

Tobacco temperature < 500 C (Cig < 900 C)

“molasses” flavorings

order of magnitude higher flow rates/puff volumes

long flow path with bubbler/humidifier (produces cool “smooth” smoke)

Tobacco smoke constituents

Smoke aerosol

Particlephase10-1000 nm(visible)

Gas phase~ 90% wt(invisible)

“tar”

condensed water

nicotine

water vaporCON2, O2, CO2

C2-C6 hydrocarbons, aldehydes, nitriles …

Illustration from: N. deNevers Pollution Control

4000+ compounds PAH, nitrosamines, metals,… contains most important carcinogens

disorder contributing agents possible enhancing agents

tobacco dependence major: nicotine

minor: secondary Nicotiana alkaloids, flavor components

acetaldehyde

cardiovascular disease major: carbon monoxide, nitrogen oxides, hydrogen cyanide, tar

minor: cadmium, zinc

nicotine, alkylating species

chronic obstructive pulmonary disease

hydrogen cyanide, volatile aldehydes, nitrogen oxides, carbon monoxide, tar

lung and larynx cancer major: PAH, NNK

minor: 210polonium, formaldehyde, acetaldehyde, butadiene, metals (Cr, Cd, Ni)

catechol, tumor promotersacetaldehydes, diet, alkylating species

oral cavity cancer major: NNN, NNK

minor: PAH

herpes simplex, irritation

esophageal cancer NNN ethanol, diet

pancreas cancer NNK, NNAL diet

Hoffmann et al, 2001; Hecht 1997

The “Hoffmann List“ of probable causative agents in cigarette smoke-related disorders

disorder contributing agents possible enhancing agents

tobacco dependence major: nicotine

minor: secondary Nicotiana alkaloids, flavor components

acetaldehyde

cardiovascular disease major: carbon monoxide, nitrogen oxides, hydrogen cyanide, tar

minor: cadmium, zinc

nicotine, alkylating species

chronic obstructive pulmonary disease

hydrogen cyanide, volatile aldehydes, nitrogen oxides, carbon monoxide, tar

lung and larynx cancer major: PAH, NNK

minor: 210polonium, formaldehyde, acetaldehyde, butadiene, metals (Cr, Cd, Ni)

catechol, tumor promotersacetaldehydes, diet, alkylating species

oral cavity cancer major: NNN, NNK

minor: PAH

herpes simplex, irritation

esophageal cancer NNN ethanol, diet

pancreas cancer NNK, NNAL diet

*Hoffmann et al, 2001; Hecht 1997

The “Hoffmann List“ of probable causative agents in cigarette smoke-related disorders

Particle size & lung dosimetry

“Ultrafine particles”Hinds, Aerosol Technology (1999)

Tobacco smoke particle (100-200 nm)

Human hair

Do waterpipes emit toxicants?

Do waterpipes emit toxicants?

What is in the smoke?

1) Find out how people smoke waterpipes

2) Program a robot to smoke the same way

3) Analyze the smoke for toxicants

Find out how people smoke Shihadeh, Antonius, Azar, BRIMC,2005

Puff topography record

Average puff topography• Volume• Duration• Frequency• Number

Field study of waterpipe users in a Beirut seaside café

Results

Puff parameter NarghileShihadeh et al 2004

CigaretteDjordjevic et al

2000

Number of puffs 171 12.1

Session smoking time, min 61 3.7

Puff volume, ml/puff 530 44.1

Puff duration, s/puff 2.6 1.5

Interpuff interval, s/cycle 17 18.5

Total volume, ml 90,600 523

A. Shihadeh et al, Biochemistry, Pharmacology, and Behavior, 79(1),75-82, 2004

Field study of 52 café smokers in Beirut mean age 21 years, 14 f/38 m

puff regimen• volume• duration• frequency• number

Ma’ssel quantity

charcoal type and application regimen

head preparation

“Beirut method” – a standard waterpipe smoking machine protocol for generating WTS

10 g ma’assel tobacco mixture load

1 easy-light charcoal disk + 0.5 added at 105th puff

18-hole aluminum perforation pattern

171 puffs, 530 ml, 2.6 s duration, 17 s IPI

the “Beirut method”

Program smoking robot & sample/analyze smoke

Shihadeh & Azar, JAM, 2006

Waterpipe smoke does contain toxicants (MS yields/unit)

NAT (ng)NNK (ng)NNN (ng)NAB (ng)

Lead, ng/10Chromium, ng/10

Arsenic, ng

Acrolein, ugAcetaldehyde, ug/10

Formaldehyde, ug

Indeno(1,2,3-cd)pyrene, ngDibenz(a,h)anthracene, ng

Benzo(a)pyrene, ng

Carbon monoxide, mgNicotine, ug/10

"Tar", mg

0 100 200 300 400 500 600 700 800 9001000

Cigarette Waterpipe (AUB-BfR)

PAHs

Aldehydes

Heavy metals

T/N/CO

TSNA

narghile cigaretteParticle mass distribution mass median dia mm 0.61 0.33Respirable mass mg/puff 0.74 0.21Nanoparticles #/puff 1.02x1011 1.11x1011

count median dia, nm 155 123

Waterpipe smoke particles are small

Waterpipe smoke particles are biologically active

Rammah et al, 2013 Toxicology Letters Volume 219, Issue 2 2013 133 - 142

In vitro effects of WTS on HEACs• Oxidative stress• Inflammation• Cell cycle arrest • Impaired vasodialation• Impaired angiogenesis

Plausible cellular mechanism for vascular diseases

Rammah et al, 2013 Toxicology Letters Volume 219, Issue 2 2013 133 - 142

Impaired angiogenesisCapillary tube formation of untreated and treated HAEC cells (mg/ml) WSC.

Waterpipe smoke particles are biologically active

Rammah et al, 2012 Toxicology Letters

Effects on lung epithelial (A549) and endothelial cells and signaling mechanisms

Plausible cellular mechanism for COPD

Hoechst staining of cells 72 hours post-treatment with WSC. Pictures were taken using a 40x oil immersion lens.

Does the user inhale toxicants?

• During a single WP use session the user inhales a large dose of toxicants known to cause tobacco-related diseases

• WTS particle size distribution is similar to cigarette smoke• WTS damages and interferes with repair mechanisms of lung

and vascular cells in culture

YES. Based on the Beirut Method:

Q. Does the Beirut Method provide a reasonable facsimile of real smoke?

RESULTS charcoal electric charcoal contribution

CO, mg 57.2 5.7 90%PAHs, ng/mg TPM

Benzo(a)pyrene 0.219 < 0.01 > 95%∑ 2- and 3-ring PAH 5.262 1.444 73%∑ 4- and 5-ring PAH 1.181 0.098-0.290a 75-92%

Charcoal(!)

Monzer, B., Sepetdjian, E., Saliba, N. and Shihadeh, A. Charcoal combustion as a source of CO and carcinogenic PAH in mainstream narghile waterpipe smok, Food and Chemical Toxicology, 2008

charcoal electrical heater temperature vs time

More charcoal than ma’ssel is consumed during a typical use session

Users continually “tune” it

REALTIME sampling in the natural environment

Katurji et al, Inh Tox, 2010

c) nicotine:tar

a) nicotine b) “tar”

d) CO

Nicotine “Tar”

CONicotine:Tar

Volume (l) Volume (l)

Volume (l)“tar” (mg)

Beirut Method

How does Beirut Method smoke compare to smoke made by people?

Katurji et al, Inh Tox, 2010

Do waterpipe users absorb toxicants?

Do waterpipe users absorb toxicants?

Contains “tar”, CO, Nicotine,PAHs, aldehydes….

What is in the smoker?

Measure toxicant levels in blood, breath, or urine.

Studies: placebo control, cigarette comparison, observational.

CO and nicotine in blood

Carboxyhemoglobin (N=31)

0

1

2

3

4

5

0 5 15 30 45

Perc

ent

Time relative to smoking onset

Plasma nicotine (N=31)

0

2

4

6

8

10

12

14

0 5 15 30 45

Time relative to smoking onset

ng/m

l

Eissenberg & Shihadeh, American Journal of Preventive Medicine, 37, 518-523, 2009.

Clinical setting, one 45 min WP session

CO and nicotine in blood: compared to cigarettes

Carboxyhemoglobin (N=31)

0

1

2

3

4

5

0 5 15 30 45

Perc

ent

Cigarette

Waterpipe

*

**

Time relative to smoking onset

*

Plasma nicotine (N=31)

0

2

4

6

8

10

12

14

0 5 15 30 45

Time relative to smoking onset

ng/m

l

*

Eissenberg & Shihadeh, American Journal of Preventive Medicine, 37, 518-523, 2009.

CO and nicotine exposure over 24 hours

Jacob et al, 2013. Cancer Epidemiol Biomarkers Prev 2013;22:765-772.

Plasma nicotine (n=13) Exhaled breath CO (n=13)

• Hospital setting, cross-over design, N=13 dual users• All day ad libitim cigarette smoking (11 cpd mean) versus 3 WTS use sessions• Measurements on day 4 of 4-day protocol.

Carcinogen exposure over 24 hours

Jacob et al, 2013. Cancer Epidemiol Biomarkers Prev 2013;22:765-772.

Urinary NNAL (TSNA biomarker) Urinary 1-HOP (PAH biomarker)

• Hospital setting, cross-over design, N=13 dual users• All day ad libitim cigarette smoking (11 cpd mean) versus 3 WTS use sessions• Measurements on day 4 of 4-day protocol.

Carcinogens in populations of smokers

Al Ali R, et al. 2013 Tob Control doi:10.1136/

Urine samples collected from population in Aleppo. Mean cigarette use 27 cigs per day, mean WTS 2 per day.

CTRL (n=28) WP (n=24) CIG (n=23)0

20

40

60

80

100

Total NNAL ng/g creatinine*, **

*, **

Do waterpipe users absorb toxicants?

YES!

• Clinical, hospital, natural environment findings consistent with one another

• Generally consistent with what we know about toxicants in smoke:

CO WP >> cigPAH WP > cigTSNA WP < cigNicotine WP ~ cig

What happens to the smoker?

Delivered nicotine is physiologically active

sympathetic activation reduced complexity

Cobb et al., Inhalation Toxicology 2012

WP smoke suppresses “Urge to smoke” after 24 hour abstinence

Rastam et al., 2011

WP smoke compromises cardiac autonomic regulation in human participants (with or without nicotine!)

0

1

2

3

4

baseline end 15 min

LF/HFTobacco

Herbal*

*

0

0.5

1

1.5

2

baseline end 15 min

SampEn Tobacco

Herbal

* *

baseline end 15 min baseline end 15 min

sympathetic activation reduced complexity

Cobb et al., Inhalation Toxicology 2012

What happens to the smoker?

WP smoke inhalation induces inflammation and oxidative stress in mice

Khabour et al 2012 Inh Tox

Inflammatory markers Oxidative stress

WP smoke inhalation associated with:

• Genotoxicity (sister chromatid exchanges): WTS > CS > nonsmokers (Khabour et al, 2011)

• Reduction in exercise capacity : VO2 1.86 vs. 1.7 l/min, pre- post (Hawari et al., 2013)

• Reduction in lung function (Hakim et al., 2011; Mohammad et al., 2008; Koseoglu et al., 2006; Kiter et al., 2000; Aydin et al. 2004; Al Mutairi et al., 2006, Al Fayez et al., 1988):

WTS compared to:Parameter a) non-smoker b) cigarette smokerFEV1 -4.04% nsFVC -1.38% nsFEV1/FVC -3.08% ns

“WPS negatively affects lung function and may be as harmful as cigarette smoking. WPS, therefore, is likely to be a cause of COPD.” – Raad et al., 2011

What happened to the smoker?

What happened to the smoker?

Waked, Khayat, Salameh, 2012

What happened to the smoker?

(Jawad et al., 2013 based on Akl et al., 2010)

“A wide range of diseases have been associated with WTS, but research in this area is relatively underdeveloped and a better evidence base is needed.” - Akl et al., 2010

Does waterpipe smoking emit toxicants into the environment?

http://firefighterparamedicstories.blogspot.com/2011_10_01_archive.html

What happens to the user?• acute physiological effects• BP, HR, inflammatory responses,

lung function…

Does waterpipe smoking emit toxicants?

exhaled mainstream (EMS)

sidestream (SS)

SS + EMS

Does the user inhale toxicants?• particle size• chemistry• biological activity

Does the user absorb toxicants?biomarkers in: blood, breath, urine

absorbed

exhaledinhaled

What happened to the smoker?• long term mortality & morbidity

Do waterpipe smoking emit toxicants?

• chamber based studies • observational studies of cafés

Daher et. al, Atmospheric Environment, 44, 8-14, 2009.

Chamber study

Waterpipe emits more nanoparticles

Daher et. al, Atmospheric Environment, 44, 8-14, 2009.

Particle concentration (particles/cm3)

WP emits more of everything measured

Daher et. al, Atmospheric Environment, 44, 8-14, 2009.

mean±95% CI waterpipe SS cigarette SS N = 12 N=9Carbon monoxide, mg 2269 ± 108 65.5 ± 5.5

PAH, ng N = 11 N = 3Total PAH 1193 ± 226 305 ± 49 Particle number emissions N = 4 N = 4ultrafine particles 5.6-99.5 nm, /1012 3.99 ± 0.60 0.639 ± 0.188total particles 5.6-560 nm, /1012 4.38 ± 0.66 1.68 ± 0.27count median diameter, nm 37.9 ± 4.1 130 ± 8 Volatile aldehydes, ug N = 6 N = 5Total aldehydes 12000 ± 1610 2954 ± 416

• Waterpipe smokers release equivalent of 2-10 cigarette smokers per hour of smoking

What about per smoker-hour?

Daher et. al, Atmospheric Environment, 44, 8-14, 2009.

Observational studies in WP cafés

* 1-Hour PM2.5 exposure Guideline-Alberta

*

Hammal et al., 2013

Observational studies in WP cafés

Zhang et al., 2013

Summary

• Delivers a large dose of toxicants

• Elicits inflammatory responses and dysfunction in human cells

• Elicits immediate physiological changes in users, including compromised lung function and autonomic function, subjective effects

• Is addictive

• Is associated with elevated markers of genetic damage and COPD

• Is associated with other long term health effects, but quality of evidence is very weak.

• Emits large quantities of toxicants in second-hand smoke

WTS…

Thanks for listening

RESULTS base case electrically heated charcoal contribution

CO, mg 57.2 5.7 90%PAHs, ng/mg TPM

Benzo(a)pyrene 0.219 < 0.01 > 95%∑ 2- and 3-ring PAH 5.262 1.444 73%∑ 4- and 5-ring PAH 1.181 0.098-0.290a 75-92%

Monzer, B., Sepetdjian, E., Saliba, N. and Shihadeh, A. Charcoal combustion as a source of CO and carcinogenic PAH in mainstream narghile waterpipe smok, Food and Chemical Toxicology, 2008

conc

entra

tion,

arb

itrar

y un

its

electrically heated, R2 < 0.02

base casecharcoal extractR2 = 0.94

charcoal grilling (R2 = 0.96, Dyremark et al, 1995)

Relative PAH concentrations measured in the mainstream smoke of the base and electrically heated conditions, and in extracts of the unburned charcoal. PAH concentrations from smoke collected from a charcoal grill by Dyremark et al (1995) shown for comparison. Correlation coefficients are shown relative to the base condition. Unburned charcoal extract, charcoal grilling smoke, and base case narghile smoke exhibit similar PAH patterns. Electrically heated condition produces a different PAH pattern.

Monzer, B., Sepetdjian, E., Saliba, N. and Shihadeh, A. Charcoal combustion as a source of CO and carcinogenic PAH in mainstream narghile waterpipe smok, Food and Chemical Toxicology, 2008

Tobacco Soex (N = 32) Baseline End 15 min Baseline End 15 minHR (1/min) 70.6 75.5* 72.2 72.8 71.5 68.1*LF (ms2) 1400 2420* 1190 1330 3570* 1610HF(ms2) 1270 1230 1140 1410 1690 1400LF/HF 1.67 2.84* 1.68 1.45 3.21* 1.55SampEn 1.63 1.48* 1.55 1.69 1.52* 1.65

Compromised cardiac autonomic regulation in human participants

Tobacco SoexTopography (N=32)

Total smoke volume (l) 31 (22) 57 (34)*Puffs drawn 95 (115) 95 (58)Mean puff volume (ml) 420 (250) 680 (330)*Mean interpuff interval (s) 45 (34) 35 (22)*Mean puff duration (s) 2.3 (0.9) 3.5 (2.0)*

Peak change from baseline plasma nicotine and CO

Nicotine (N=32) 7.8 (8.0) 0.0 (0.0)*CO% (N=29) 11.8 (7.7) 30.9 (19.0)*

WP emits more of everything measured

Daher et. al, Atmospheric Environment, 44, 8-14, 2009.

mean±95% CI waterpipe SS cigarette SS N = 12 N=9Carbon monoxide, mg 2269 ± 108 65.5 ± 5.5

PAH, ng N = 11 N = 3Total PAH 1193 ± 226 305 ± 49 Particle number emissions N = 4 N = 4ultrafine particles 5.6-99.5 nm, /1012 3.99 ± 0.60 0.639 ± 0.188total particles 5.6-560 nm, /1012 4.38 ± 0.66 1.68 ± 0.27count median diameter, nm 37.9 ± 4.1 130 ± 8 Volatile aldehydes, ug N = 6 N = 5Total aldehydes 12000 ± 1610 2954 ± 416

Mass balance

𝑚𝑒=𝑉 [𝐶 ( 𝑡𝑠 )−𝐶 (0 ) ]+∫0

𝑡 𝑠

𝐶 (𝑡 )𝑄𝑑𝑡+𝑚𝑑

change of suspended mass within CV

For any toxicant…

advected mass

wall losses

Sidestream emissions

Daher et. al, Atmospheric Environment, 44, 8-14, 2009.

mean±95% CI waterpipe SS cigarette SS N = 12 N=9Carbon monoxide, mg 2269 ± 108 65.5 ± 5.5

PAH, ng N = 11 N = 3Total PAH 1193 ± 226 305 ± 49

Particle number emissions N = 4 N = 4

ultrafine particles 5.6-99.5 nm, /1012 3.99 ± 0.60 0.639 ± 0.188

total particles 5.6-560 nm, /1012 4.38 ± 0.66 1.68 ± 0.27count median diameter, nm 37.9 ± 4.1 130 ± 8

Volatile aldehydes, ug N = 6 N = 5Total aldehydes 12000 ± 1610 2954 ± 416

SS nanoparticle emissions: WP vs CIG

Daher et. al, Atmospheric Environment, 44, 8-14, 2009.

Particle concentration (particles/cm3)

Is robot-generated smoke realistic?

y = 0.018x + 1.470R² = 0.622

0

4

8

12

16

0 200 400 600 800

Bloo

d CO

(CO

Hb, %

)

Smoking machine CO (mg)

Toxicants in robot-generated smoke track blood level exposure in human participants

Carbon monoxide

Shihadeh & Eissenberg, CEBP, 2011

P<0.01R2>0.62

Schubert et al., 2012

Environ Mol Mutagen. 2011 Apr;52(3):224-8. doi: 10.1002/em.20601. Epub 2010 Aug 25. Assessment of genotoxicity of waterpipe and cigarette smoking in lymphocytes using the sister-chromatid exchange assay: a comparative study.Khabour OF, Alsatari ES, Azab M, Alzoubi KH, Sadiq MF.

AbstractTobacco smoking is a major world health problem. Recently, waterpipe smoking has become more popular in many countries. Although the genotoxicity associated with cigarette smoking has been extensively investigated, studies evaluating such toxicity in waterpipe users are still lacking. In this study, we examined the genotoxicity of waterpipe smoking in lymphocytes compared with the genotoxicity of cigarette smoking. Genotoxicity was evaluated using the sister chromatid exchanges (SCEs) assay. Fifty waterpipe smokers and 18 healthy nonsmokers participated in this study. Additionally, 18 heavy cigarette smokers (CS) were recruited for comparison. The results show that waterpipe smoking and cigarette smoking significantly increase the frequencies of SCEs (P < 0.01) compared with those of nonsmokers, indicating the genotoxic effect of tobacco smoking. In addition, frequencies of SCEs were significantly higher among waterpipe smokers compared with CS (P < 0.01), indicating that waterpipe smoking is more genotoxic than cigarette smoking. Moreover, the frequency of SCEs increased with the extent of waterpipe use. In conclusion, waterpipe smoking is genotoxic to lymphocytes and the magnitude of its genotoxicity is higher than that induced by regular cigarette smoking.