Influence of Phthalates on Cytokine Production in Monocytes and Macrophages: A Systematic Review of Experimental Trials

<div><p>Background</p><p>Phthalates are a group of endocrine disrupting chemicals suspected to influence the immune system. The aim of this systematic review is to summarise the present knowledge on the influence of phthalates on monocyte and macrophage production and secretion of cytokines, an influence which could affect both pro- and anti-inflammatory abilities of these cells.</p><p>Strategy and Results</p><p>A systematic search was performed in Medline, Embase and Toxline in June 2013, last updated 3rd of August 2014. Criteria used to select studies were described and published beforehand online on Prospero (<a href="http://www.crd.york.ac.uk/NIHR_PROSPERO" target="_blank">http://www.crd.york.ac.uk/NIHR_PROSPERO</a>, registration number CRD42013004236). <i>In vivo</i>, <i>ex vivo</i> and <i>in vitro</i> studies investigating the influence of phthalates on cytokine mRNA expression and cytokine secretion in animals and humans were included. A total of 11 reports, containing 12 studies, were found eligible for inclusion. In these, a total of four different phthalate diesters, six primary metabolites (phthalate monoesters) and seven different cytokines were investigated. Though all studies varied greatly in study design and species sources, four out of five studies that investigated di-2-ethylhexyl phthalate found an increased tumour necrosis factor-α secretion/production from monocytes or macrophages. A summary of cytokine measurements was not possible since few studies were comparable in study design and due to insufficient reporting of raw data for most of the included studies.</p><p>Conclusion</p><p>Results from this review have suggested that at least one phthalate (di-2-ethylhexyl phthalate) has the ability to enhance tumour necrosis factor-α production/secretion from monocytes/macrophages <i>in vitro</i>, but also observed <i>ex vivo</i>. Influence of other phthalates on other cytokines has only been investigated in few studies. Thus, <i>in vitro</i> studies on primary human monocytes/macrophages as well as more <i>in vivo</i> studies are needed to confirm or dispute these findings.</p></div

Primary and secondary outcomes from individual studies.

<p>Footnote: BBzP: Butylbenzyl phthalate, DEHP: Di-2-ethylhexyl phthalate, DiNP: Di-iso-nonyl phthalate, DnBP: Di-n-butyl phthalate, IL: Interleukin, LDH: lactate dehydrogenase, MEHP: Mono-(2-ethylhexyl) phthalate, ND: not done, PI: Propidium Iodide, RAW 264 cell line: mouse leukemic monocyte-macrophage cell line, SD: standard deviation, SEM: standard error of the mean, THP-1 cell line: acute monocytic cell line, TNF: Tumour necrosis factor.</p><p>Primary and secondary outcomes from individual studies.</p

Study characteristics.

<p>Footnote:</p><p>*: macrophage/monocyte content not confirmed, but cells stimulated with LPS.</p><p>†: macrophage content confirmed by flow cytometry.</p><p>BBzP: Butylbenzyl phthalate, DEHP: Di-2-ethylhexyl phthalate, DiNP: Di-iso-nonyl phthalate, DnBP: Di-n-butyl phthalate, IL: interleukin, MBzP: Mono-benzyl phthalate, MEHP: Mono-(2-ethylhexyl) phthalate, MiDP: Mono-iso-decyl phthalate, MiNP: Mono-iso-nonyl phthalate, MnBP: Mono-n-butyl phthalate, MOP: Mono-n-octyl phthalate, NRCT: Non randomised controlled trial, PMA: Phorbol 12- myristate 13-acetate, RAW 264 cell line: mouse leukemic monocyte-macrophage cell line, RCT: Randomised controlled trial, THP-1 cell line: acute monocytic cell line, TNF: tumour necrosis factor.</p><p>Study characteristics.</p

Monoester concentrations (mean ± SD) in cell culture supernatants with the highest added diester-concentrations (1 to 100 μM).

<p>N = DEP: one culture in duplicates; DnBP and DEHP: two cultures in single determination except for 100 μM DEHP, with only one culture in single determination due to removal of a single outlier. Grey dashed lines indicate the expected monoester concentration if all diester had been metabolised. DEP: di-ethyl phthalate. DnBP: di-n-butyl phthalate. DEHP: di-2-ethylhexyl phthalate.</p

Levels of outcome variables in un- and TSH-stimulated controls.

<p>Experiments were conducted in duplicates or triplicates and exposure time was 72 h. Log10-transformed data was used in statistical analysis. cAMP: 3'-5'-cyclic adenosine monophosphate. IL: interleukin. NIS: sodium iodine symporter. Tg: thyroglobulin. TPO: thyroid peroxidase. TSHr: thyroid stimulating hormone receptor.</p

Time study of phthalate diester metabolism.

<p>The measured content of MEP, MnBP and MEHP are expressed as mean ± SD at time points 0 to 72 hours after phthalate diester addition (0.1 μM DEP, DnBP or DEHP) to TSH-stimulated cell cultures (n = three cultures in single determination). The monoester concentrations are corrected for background contamination. MEP: mono-ethyl phthalate. MnBP: mono-n-butyl phthalate. MEHP: mono-2-ethylhexyl phthalate.</p