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# Chlordecone (kepone) induces mitochondrial dysfunction in human cardiac tissue

Alexia Fundere, Marie-Daniela Dubois, Magalie Vatin Inamo, Fatima Radouani, Prisca Jalta, Dabor Resiere, Remi Neviere

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## To cite this version:

Alexia Fundere, Marie-Daniela Dubois, Magalie Vatin Inamo, Fatima Radouani, Prisca Jalta, et al. Chlordecone (kepone) induces mitochondrial dysfunction in human cardiac tissue. Mitochondrion, 2025, 84, 10.1016/j.mito.2025.102058. hal-05209490

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Original Research Article

# Chlordecone (kepone) induces mitochondrial dysfunction in human cardiac tissue

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Alexia FUNDERE<sup>a,b</sup>, Marie-Daniela DUBOIS<sup>a</sup>, Magalie VATIN INAMO<sup>a,b</sup>, Fatima RADOUANI<sup>a,c</sup>, Prisca JALTA<sup>a,c</sup>, Dabor RESIERE<sup>a,d</sup>, Remi NEVIERE<sup>a,e,*</sup>

ARTICLE INFO

Keywords:

Chlordecone

Cardiac tissue

Human heart

Mitochondria, oxygen consumption

ABSTRACT

Chlordecone exposure in humans has been associated with increased incidence of prostate cancer, impaired fertility, and fetal/perinatal abnormalities while experiment rodent studies suggest that chlordecone can inhibit magnesium-ATPase, little is known about its mitochondrial toxicity in humans. Our objective was to test whether chlordecone would induce mitochondrial dysfunction in human cardiac cells in *ex vivo* heart preparations. Biopsies of human atrial tissue were obtained during cannulation for cardiopulmonary bypass from patients who were undergoing programmed cardiac surgery for coronary artery bypass. Cardiac preparations were incubated with vehicle or chlordecone (5 nM and 50 nM) for 24 hr followed by mitochondrial high-resolution oxygraphy studies. Compared with vehicle, chlordecone cardiac exposure at the concentrations of 5 nM and 50 nM impaired mitochondrial respiratory rates. Chlordecone concentrations of 5 nM and 50 nM similarly increased state 2 respiration rate and maximal respiration capacity with no change of state 3 (ADP) respiration rate, which suggests the uncoupling of between mitochondrial oxidative phosphorylation and electron transport through the respiratory chain complexes. In conclusion, our study suggests that chlordecone at clinically relevant concentration impairs mitochondrial function leading to uncoupling, which may induce abnormal cardiac cell responses, including aberrant calcium handling and oxidative stress.

1. Introduction

Chlordecone, also known as kepone, is an organochlorine insecticide that has been used to eradicate borers of the banana roots in the French West Indies, Guadeloupe and Martinique. Chlordecone has been used from 1973 to 1993, while it has been prohibited in the USA in 1976 (Benoit et al., 2020; Inserm, 2022; Nedellec et al., 2016; Nedellec et al., 2016). Chlordecone was available till 1981 in the French West Indies and was eventually banned in 1993. Due to its persistent nature in the environment and ecosystem, chlordecone has led to the insidious poisoning of the population for more than thirty years (Benoit et al., 2020; Inserm, 2022; Nedellec et al., 2016; Nedellec et al., 2016).

As a part of the endocrine disruptor effects, chlordecone displays well recognized estrogenic and progestagenic properties. The estrogenic

and progestagenic-like actions of chlordecone have been clearly established both *in vivo* and *in vitro* experiments. Chlordecone binds to estrogen receptors α (ERα) and β (ERβ), which induce ERα agonist and Erβ antagonist activities. In addition to the nuclear ERs interaction, chlordecone may also stimulate alternative estrogen signaling pathways and bind to receptors involved in steroid homeostasis (Inserm, 2022; Baratzhanova et al., 2024). *In vitro* tests also revealed evidence that chlordecone may act as a agonistic and/or antagonistic molecule on androgen receptor (AR) and cytochrome P450 19A (CYP19A) (Inserm, 2022; Baratzhanova et al., 2024). In line with these endocrine disruptor effects, blood concentrations of chlordecone above 0.5–1 µg per liter of plasma have been associated with increased incidence of prostate cancer, impaired fertility, as well as fetal/perinatal and psychomotor development abnormalities in the Caribbean population (Brureau et al.,

<footnote><sup>a</sup> Cardiovascular Research Team (UR5_3 PC2E), University of the French West Indies (Université des Antilles), 97200 Fort de France, France</footnote>

<footnote><sup>b</sup> Department of Biology, Faculty of Natural and Exact Sciences, University of the French West Indies (Université des Antilles), 97110 Pointe-à-Pitre, France</footnote>

<footnote><sup>c</sup> Department of Research, University Hospital of Martinique (CHU Martinique), 97261 Fort-de-France, France</footnote>

<footnote><sup>d</sup> Department of Toxicology and Critical Care Medicine, University Hospital of Martinique (CHU Martinique), 97261 Fort-de-France, France</footnote>

<footnote><sup>e</sup> Department of Clinical Physiology, University Hospital of Martinique (CHU Martinique), 97261 Fort-de-France, France</footnote>

<footnote>* Corresponding author at: Cardiovascular Research Team (UR5_3 PC2E), University of the French West Indies (Université des Antilles) and Department of Clinical Physiology, CHU Martinique (University Hospital of Martinique), 97261 Fort de France, France.</footnote>

<footnote>E-mail address: remi.neviere@chu-martinique.fr (R. NEVIERE).</footnote>
2020; Costet et al., 2015; Hervé et al., 2016; Multigner et al., 2010; Multigner et al., 2008).

Besides, chlordecone may induce alterations of cellular calcium ho-
meostasis (End et al., 1979; Mishra et al., 1980; Agarwal and Mehendale,
1984; Kodavanti et al., 1990; Kodavanti et al., 1991). Previous studies
suggest that chlordecone inhibits calcium (Ca²⁺) transient by altering
Ca²⁺-ATPase, impairing sarcoplasmic reticulum Ca²⁺ uptake and dis-
rupting Ca²⁺ storage (Mishra et al., 1980; Kodavanti et al., 1990).
Chlordecone can induce perturbation of the sodium (Na⁺)-potassium
(K⁺)-ATPase pump (Folmar, 1978; Jinna et al., 1989; Maier and Costa,
1990), which may result in cardiac cells of sarcolemmal sodium Na⁺–
Ca²⁺ exchanger (NCX) dysfunction and elevated diastolic calcium con-
centration in the cytosol. Chlordecone may also inhibit magnesium
(Mg²⁺)-ATPase and disrupt mitochondria ultrastruture and energy
production (End et al., 1979; Folmar, 1978; Atwal, 1973; Carmines
et al., 1979; Desaiah et al., 1977; Desaiah et al., 1977; Desaiah, 1981;
Manring and Moreland, 1981; Soileau and Moreland, 1983). Of note,
chlordecone toxicity was evaluated using micromolar concentrations far
above clinically relevant levels.

While chlordecone can provoke malfunction of myocardial sarco-
plasmic reticulum Ca²⁺-ATPase pump and sarcolemma exchanger
(Mishra et al., 1980; Kodavanti et al., 1990; Folmar, 1978; Jinna et al.,
1989; Maier and Costa, 1990), no adverse cardiac effects have ever been
reported in the numerous animal toxicological studies that have inves-
tigated the health effects of chlordecone exposure (Inserm, 2022;
Nedellec et al., 2016; Nedellec et al., 2016). Likewise, no cardiac com-
plications, including an abnormal electrocardiogram pattern, were
noted, despite detailed and prolonged clinical monitoring of workers
exposed to elevated doses of chlordecone at the Hopewell factory (Vir-
ginia, US) (Inserm, 2022; Nedellec et al., 2016; Nedellec et al., 2016;
Cannon et al., 1978). However, a recent study reported that adult male
rats ingested with daily-doses of diluted chlordecone in their quotidian
water display abnormal myocardial function evaluated by echocardi-
ography (Fundere et al., 2024). In addition, this study suggests that
chlordecone ingestion is associated with abnormal cardiac remodeling
along with increased expressions of proinflammatory and fibrosis
markers (Fundere et al., 2024). Increased atrial arrhythmia vulnerability
induced by chlordecone exposure resulted in profound changes of
mRNA expression of potassium and sodium channels (Kcnq1, Scn5a),
which are central players of enhanced vulnerability to atrial fibrillation
(Fundere et al., 2024). Interestingly, while food intake was increased,
chlordecone-treated rats displayed major body weight reduction, sug-
gesting increased basal metabolism rate and mitochondrial oxygen
consumption when obvious spontaneous physical activity is excluded
(Fundere et al., 2024; Williams et al., 1992; Klingensmith and Mehen-
dale, 1982). Such abnormal metabolic features may be linked to the
uncoupling of mitochondrial oxidative phosphorylation (ATP) from the
oxygen consumption cycle induced by chlordecone (End et al., 1979;
Folmar, 1978; Atwal, 1973; Carmines et al., 1979; Desaiah et al., 1977;
Desaiah et al., 1977; Desaiah, 1981; Manring and Moreland, 1981).

In addition to abnormal ionic channel machinery, mitochondrial
dysfunction is a central pathological process involved in human atrial
fibrillation (Montaigne et al., 2013; Liu et al., 2021). Dysfunctional
mitochondria can generate oxygen and nitrogen free radicals that can
oxidize several intracellular targets, such as kinases, sodium and calcium
channels, and mitochondrial DNA. Such mitochondrial signaling can
directly impair cardiomyocyte excitability and favor functional reen-
trant circuits of atrial fibrillation (Montaigne et al., 2013; Liu et al.,
2021; Liu et al., 2022). Mitochondrial dysfunction further disrupts
cellular energy metabolism and cellular stress signal transduction,
which exacerbate cardiomyocyte stress responses and ultimately pro-
mote atrial fibrillation occurrence (Liu et al., 2021; Liu et al., 2022). It is
hence important to test whether chlordecone may promote mitochon-
drial dysfunction in human atrial heart that would provide explanation
for increased atrial arrhythmia vulnerability elicited by chlordecone in
the experimental model.

Of note, there is no information about the role of chlordecone on
mitochondrial respiration in the human heart. The objective of the study
was to test whether chlordecone would induce mitochondrial dysfunc-
tion in *ex vivo* human heart preparations.

## 2. Materials and methods

2.1. *Ethics Statement*

The study was conducted following the recommendations of the
criteria edited by the amended Declaration of Helsinki (https://www.
wma.net/policies-post/wma-declaration-of-helsinki/). Informed con-
sent for the processing of personal data and tissue collection for scien-
tific research purposes was obtained in each patient. The French medical
ethics committee (CPP: 19-MART-01) further approved the study
design.

## 2.2. *Chemicals*

Chlordecone, chemicals for BIOPS buffer, saponin, L-glutamate, L-
malate, rotenone, antimycin-A, cytochrome c, N,N,N',N' -tetramethyl-
p-phenylenediamine dihydrochloride (TMPD) and ascorbate (asc) were
purchased from Sigma Aldrich (St. Quentin Fallavier, France). MiRO
buffer was obtained from Oroboros (Innsbruck, Austria). Chlordecone
was dissolved in ethanol. The final concentration of the solvent was held
constant at 0.01 % in all assays, including controls.

3. Research design

After a 24 hr storage period, mitochondrial respiration was measured
in permeabilized cardiac fiber preparations. The 24 hr stability of
mitochondrial respiration of *ex vivo* human cardiac trabeculae has been
previously proven in our laboratory (Cano-Sanchez et al., 2022). Study
design compared the effects of 24 hr incubation of human cardiac
myofibers in the presence or not of chlordecone at the final concentra-
tions of 5 nM (2.45 µg/L) and 50 nM (24.5 µg/L). The study protocol is
summarized Fig. 1.

3.1. *Human cardiac tissue*

Patient's main characteristics are summarized Table 1. Right atrial
appendages were obtained during cardiopulmonary bypass from pa-
tients who were undergoing programmed surgery for coronary artery
bypass. All patients were anaesthetized with administration of volatile
anesthetic gas or propofol, sufentanil, and skeletal muscle relaxant.
Patients with medical history arrhythmia and those with poor left ven-
tricular ejection fraction (< 55 %) or right coronary stenosis were
excluded from the study. Right atrial appendages obtained at the time of
cannulation for cardiopulmonary bypass and immediately placed in ice-
cold Krebs-Henseleit buffer (KHB) for analysis. Atrial appendages were
carefully prepared under stereomicroscopic control in ice-cold relaxing
and biopsy preservation solution BIOPS as we have previously described
(Montaigne et al., 2013; Cano-Sanchez et al., 2022). Fibers were sepa-
rated with needles and forceps and then transferred to Eppendorf tube
containing 2 ml of oxygenated BIOPS buffer containing 2.77 mM
CaK₂EGTA, 7.23 mM K₂EGTA, 6.56 mM MgCl₂, 0.5 mM dithiothreitol,
50 mM K-MES, 20 mM imidazole, 20 mM taurine, 5.3 mM Na₂ATP, 15
mM phosphocreatine, pH 7.1, at 37 °C.

3.2. *High-resolution respirometry*

At the end of the 24 hr incubation time, fibers (5 mg) were trans-
ferred into 2 mL tubes filled of fresh ice-cold BIOPS relaxing solution and
20 µL of saponin. Saponin was prepared freshly from a stock solution 5
mg/mL are added to the 2 mL tube to obtain a final saponin concen-
tration of 50 µM. Cardiac fibers were maintained for 30 min with the
<figure><img src="image_6.png" /><figcaption>Fig. 1. Study protocol.</figcaption></figure>

**Table 1**

<table><caption>Patients' demography and medical history (N = 5).</caption><tbody><tr><td>Age (year)</td><td>62 ± 7</td></tr><tr><td>Gender (Male) n (%)</td><td>5 (100)</td></tr><tr><td>BMI (kg/m²)</td><td>30 ± 4</td></tr><tr><td>Systolic blood pressure (mmHg)</td><td>135 ± 8</td></tr><tr><td>Diastolic blood pressure (mmHg)</td><td>77 ± 6</td></tr><tr><td><strong>Comorbidities</strong></td><td></td></tr><tr><td>Hypertension n (%)</td><td>2 (40)</td></tr><tr><td>Diabetes n (%)</td><td>2 (40)</td></tr><tr><td>Dyslipidemia n (%)</td><td>3 (60)</td></tr><tr><td>Active smoker n (%)</td><td>2 (40)</td></tr><tr><td><strong>Laboratory data</strong></td><td></td></tr><tr><td>Creatinine (mg/L)</td><td>9.8 ± 3.5</td></tr><tr><td>Glycated hemoglobin HbA1c (%)</td><td>5.8 ± 0.6</td></tr><tr><td>HDL (g/L)</td><td>0.39 ± 0.12</td></tr><tr><td>TG (g/L)</td><td>1.4 ± 0.4</td></tr><tr><td><strong>Preoperative echocardiographic data</strong></td><td></td></tr><tr><td>LVEF (%)</td><td>56 ± 8</td></tr><tr><td><strong>Preoperative medication</strong></td><td></td></tr><tr><td>Statins n (%)</td><td>5 (100)</td></tr><tr><td>Diuretics n (%)</td><td>0 (0)</td></tr><tr><td>ACE-inhibitors or ARBs n (%)</td><td>2 (40)</td></tr><tr><td>Beta-blockers n (%)</td><td>5 (100)</td></tr><tr><td>Antiplatelet agent n (%)</td><td>5 (100)</td></tr><tr><td>Oral anticoagulant n (%)</td><td>0 (0)</td></tr><tr><td>Insulin n (%)</td><td>0 (0)</td></tr><tr><td>Metformin n (%)</td><td>0 (0)</td></tr><tr><td>Time on bypass (min)</td><td>89 ± 37</td></tr></tbody></table>

Continuous variables are given as mean ± SD and categorical data are ables as absolute values with percentages. BMI, body mass index; HDL, high density lipoprotein; TG, triglycerides; LVEF, left ventricular ejection fraction; ACE, angiotensin converting enzyme; ARBs, angiotensin receptor blockers.

saponin solution, and then washed for 5 min three times in ice-cold respiration medium MiRO₅ (Oroboros, Innsbruck, Austria). MiRO₅ is the respiratory buffer that contains 110 mM sucrose, 20 mM HEPES, 10 mM KH₂PO₄, 20 mM taurine, 3 mM MgCl₂ 6 H₂O, 60 mM MES-K, 0.5 mM EGTA and 0.1 % bovine serum albumin; pH 7.1. Respiration rates were evaluated using a high resolution oxygraphy (O₂k oxygraph,

Oroboros, Innsbruck, Austria). Mitochondrial respiratory rates were expressed in picomoles O₂ per second per milligram wet weight. Data acquisition and analysis were processed by the Datlab4 software (Oroboros, Innsbruck, Austria).

### 3.2.1. Mitochondrial respiration rates

Substrate and/or inhibitors for respiratory experiments were added within MiRO₅ buffer in a step-by-step manner using micro syringes. Chemical agents were sequentially prepared according to Oroboros manufacturer data sheet and administered as previously described (Montaigne et al., 2013; Cano-Sanchez et al., 2022; Kuznetsov et al., 2008). Respiration rates were measured as described below:

- Respiration rate in the presence of 10 mM L-glutamate and 2 mM L-malate dissolved in H₂O is related to complex I-dependent state 2 respiration (V₀). L-glutamate and L-malate activate the Krebs cycle enzyme malate dehydrogenase providing NADH to complex I of the respiratory chain.

- Phosphorylation-stimulated respiration rate (Vₗlut+mal) was determined by the adding of 5 mM ADP to the V₀ medium. Vₗlut+mal rate is related to complex I-dependent state 3 respiration as ADP to F1-F0 ATP synthase.

- Complex I inhibitor rotenone (0.5 µM) dissolved in ethanol was added to the medium allowing the determination of complex II-dependent uncoupled state of respiration (Vₚrot).

- Respiration rate in presence of complex III inhibitor antimycin-A (2.5 µM) dissolved in ethanol and added to the Vₚrot medium is related to O₂ consumption independent of the respiratory-chain (Vₐₐ). In this case, electron supply to complex IV, and all respiratory chain complexes are stopped.

- Maximal respiration rate related to complex IV-dependent uncoupled state of respiration (Vₐₐₓ) was then determined. Firstly, O₂ consumption rate was evaluated after the addition of 2 mM ascorbate (asc) and 0.5 mM N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride (TMPD) dissolved in H₂O into the Vₐₐ medium (VₗTMPD-asc). TMPD was used as an artificial redox activator that boosts electron transfer from ascorbate to cytochrome c. The auto-oxidation of TMPD-ascorbate was then determined as the O₂ utilization (O₂ consumption) in presence of 1 mM KCN added to the VₗTMPD-asc medium. The difference VₗTMPD-asc minus VₚKN was calculated and related to complex IV-dependent uncoupled state of respiration (Vₐₐₓ). In this case, Vₐₐₓ estimates complex IV-related maximum respiration rate.
- At the end of each procedure, zero O₂ content calibration was performed with dithionite.

### 3.2.2. Intactness of mitochondrial outer membrane

We evaluated the intactness of mitochondrial outer membrane indirectly by the use of exogenous cytochrome c. When exogenous cytochrome c (10 µM cytochrome c dissolved in ethanol (horse heart cytochrome c, Sigma Aldrich, France) ($V_{cyt-c}$) respiration rate may increase whether mitochondrial outer membrane permeability is impaired. The $V_{cyt-c}/V_{glut+mal}$ ratio is hence a permeability index of the outer mitochondrial membrane (Kuznetsov et al., 2008).

## 3.3. Statistics

A complete descriptive analysis was performed using SPSS for Windows software, version 24.0 (SPSS Paris La-Defense, France). For all descriptive and inferential analyzes, the assumption of normal data distribution was analyzed by the Shapiro-Wilk test. Mean and standard deviation were reported for continuous normally distributed data, whereas median, interquartile, and min-max ranges were described for continuous non-normally distributed data. Categorical variables were presented as frequencies or percentages. The following tests were used for group comparisons for quantitative continuous variables: Student's t-test (parametric test) or Wilcoxon-Mann-Whitney test (non-parametric test). All inferential analyses were performed by means of a 2-sided test, with a level of significance of 5%.

# 4. Results

After 24 hr incubation with chlordecone or solvent (vehicle), measurements of mitochondrial respiration rates of saponin-permeabilized cardiac fibers were performed. Main results of mitochondrial respiration are summarized Figs. 2-5. Mitochondrial respiration state 2 ($V_0$) rates were higher in chlordecone-treated cardiac fibers compared with controls, while ADP-coupled mitochondrial respiration state 3 ($V_{glut+mal}$) rates were similar, suggesting uncoupling (Fig. 2).

Respiration rates in the presence of rotenone ($V_{rot}$) are displayed Fig. 3.

Respiration rates in the presence of Antimycin A ($V_{AA}$) are displayed Fig. 4.

Maximum mitochondrial respiration ($V_{Cox}$) rates were higher in chlordecone-treated cardiac fibers compared with controls (Fig. 5).

Mitochondrial outer membrane intactness was indirectly evaluated by measuring reduction of exogenous cytochrome c (Cyt c) (Kuznetsov et al., 2008). In mitochondria, Cyt c is displayed at the mitochondrial intermembrane/inter-cristae spaces, where it acts as an electron shuttle driving the respiratory chain complex processes. Whether mitochondrial membrane permeability is impaired, more cytochrome c can enter into

<figure><img src="image_7.png" /><figcaption>**Fig. 2.** Respiration rate in the presence of glutamate-malate. Panel A shows complex I-dependent state 2 respiration $V_0$ (state 2). Panel B shows ADP-stimulated respiration rate $V_{glut+mal}$ (state 3). Results are displayed as box plot with median, min to max, and single data points. N = 5, ns indicates non-significant, * indicates p < 0.05 versus vehicle.</figcaption></figure>

<figure><img src="image_8.png" /><figcaption>**Fig. 3.** Respiration rate in the presence of rotenone ($V_{rot}$). Results are displayed as box plot with median, min to max, and single data points. N = 5, ns indicates non-significant.</figcaption></figure>

<figure><img src="image_9.png" /><figcaption>**Fig. 4.** Respiration rate in the presence of Antimycin A ($V_{AA}$). Results are displayed as box plot with median, min to max, and single data points. N = 5, ns indicates non-significant.</figcaption></figure>

<figure><img src="image_10.png" /></figure>
<figure><img src="image_11.png" /><figcaption>**Fig. 5.** Maximal respiration rate in the presence of TMPD and ascorbate (V<sub>COx</sub>). Results are displayed as box plot with median, min to max, and single data points. N = 5, * indicates p < 0.05 versus vehicle.</figcaption></figure>

the mitochondria and stimulates mitochondrial respiration. Typically, an increase of mitochondrial respiration up to 8 % suggests impaired mitochondrial membrane permeability (Kuznetsov et al., 2008). In our study, increases of V<sub>glut+mal</sub> complex I-dependent (state 3) respiration in the presence of cytochrome c 10 µM was 3.2 ± 0.5 % in controls versus 4.0 ± 1.1 % in chlordecone-treated cardiac fibers, suggesting mitochondrial membrane intactness.

## 5. Discussion

To the best of our knowledge, our study is the first to suggest that chlordecone exposure of human cardiac cells using clinically relevant concentrations impairs mitochondrial function leading to uncoupling, a condition in which electron transport along respiratory chain complexes is disconnected from the phosphorylation of ADP to ATP (Murphy, 2009; Demine et al., 2019). In our study, nanomolar concentrations of chlordecone were used to expose human heart preparation with clinically relevant concentrations. Tested chlordecone concentrations of 5 nM and 50 nM were within the range of the clinical toxicological reference value of chlordecone (0.4 µg/L) in the plasma. Our findings are consistent with previous studies showing that chlordecone might generate the uncoupling of mitochondrial oxidative phosphorylation (ATP) from the oxygen consumption cycle. Both *in vitro* and *in vivo* experiments in rodents have shown that chlordecone at micromolar concentrations can disrupt mitochondria ultrastructure, reduce mitochondrial Mg<sup>2+</sup>-ATPase activity and impair mitochondrial respiration (End et al., 1979; Folmar, 1978; Jinna et al., 1989; Atwal, 1973; Carmines et al., 1979; Desaiah et al., 1977; Desaiah et al., 1977; Desaiah, 1981; Manring and Moreland, 1981; Soileau and Moreland, 1983). Another study confirmed that chlordecone used at nanomolar concentrations found in the drinking water and plasma levels of exposed population in the French West Indies (Guadeloupe and Martinique) also increase mitochondrial uncoupled respiration in human umbilical vein endothelial cells (Alibrahim et al., 2018).

As a consequence of mild-uncoupling, mitochondrial membrane potential is dissipated, allowing for the temporarily reduction of calcium influx and attenuate oxygen free radical generate by the mitochondria (Hirschenson et al., 2022). As a different concept, uncoupling of respiration due to proton leak can elicit excessive mitochondrial ROS production and mitochondrial calcium overload (Wallace and Starkov, 2000). Mitochondrial uncoupling has been previously implicated in cardiovascular diseases through mechanisms including impaired mitochondrial ATP synthesis, cardiac excitation-contraction coupling and

mitochondrial ROS. It may be hence speculated that uncoupling properties of chlordecone observed in our human heart experiments is associated with increased mitochondrial ROS. Considering mitochondrial dysfunction as central process involved in human atrial fibrillation and heart remodeling, mitochondrial uncoupling observed in our human cardiac preparations may explain the recent reported findings that chlordecone promote the occurrence of cardiac arrhythmias and myocardial remodeling (Fundere et al., 2024).

Mitochondrial uncoupling reported in experimental studies (End et al., 1979; Folmar, 1978; Atwal, 1973; Carmines et al., 1979; Desaiah et al., 1977; Desaiah et al., 1977; Desaiah, 1981; Manring and Moreland, 1981) and suggested by our results in human cardiac preparation may represent a potential mechanism involved in the unexplained weight loss observed in workers (Cannon et al., 1978) and rodents (Fundere et al., 2024; Williams et al., 1992; Klingensmith and Mehendale, 1982) exposed to chlordecone. It appears that chlordecone can induce weight loss as a consequence of body fat store depletion and altered energy balance, in a scenario similarly to the well-known 2,4-dinitrophenol (DNP) uncoupler (Grundlingh et al., 2011). DNP is an uncoupling agent of the oxidative phosphorylation that has been used in the early 1930 s for weight loss. The use of DNP was quickly banned by the U.S. Food and Drug Administration (FDA) because of its severe cardiovascular. As chlordecone stimulates mitochondrial respiration in a manner similar to uncouplers of oxidative phosphorylation, the effects of chlordecone and 2,4-dinitrophenol (DNP) have been previously compared to chlordecone (Desaiah et al., 1977; Desaiah et al., 1977; Desaiah, 1981). While chlordecone was found a more potent uncoupler than DNP, chlordecone strongly inhibited the effect of DNP suggesting a similar mitochondrial site of action. It is hence proposed that mitochondrial uncoupling effect of chlordecone may increase basal metabolic rate and elicit weight loss. A schematic to summarize the proposed mechanical cascade from chlordecone exposure to mitochondrial dysfunction and potential consequences on cardiac function is displayed in Fig. 6.

This study has several limitations. Firstly, explanation for why chlordecone at two different concentrations induced similar results on mitochondrial uncoupling is not readily available. This unexpected result may be related to the Q-channel structure of mammalian mitochondrial complex I, which may function in all-or-nothing mode with a “closed” state, with a tight and sealed channel, and an “open” state with more relaxed and disordered loops (Pereira et al., 2023). Secondly, we acknowledge that a very small number of biopsies were evaluated in our study. Likewise, samples were of small size limiting the possibility to explore complementary mechanisms associated with mitochondrial dysfunction such as oxidative stress evaluation and mitochondrial signaling. Also, the range of chlordecone concentration was limited to two nanomolar concentrations. The limitation was mainly due to the restricted access to cardiac surgery with extracorporeal circulation bypass in our Caribbean territory. Of note, open heart surgery facility is restricted to the University Hospital of Martinique in the Caribbean basin. Thirdly, mitochondrial respiration was investigated in cardiac fibers obtained from patients undergoing coronary artery bypass surgery, who display coronary artery diseases caused by a variety of associated factors, such as dyslipidemia smoking and diabetes. In order to limit the effects of coronary atherosclerosis, patients with right artery stenosis were excluded, which avoid major hypoxia of the right atrial tissue and ensure the integrity of mitochondria used for respiration evaluation. It is also to note that the same biopsy was split in two part and used for control and chlordecone conditions. This protocol may reduce the confounding effects of mitochondrial preparation heterogeneity between experimental conditions. The control condition and the chlordecone exposure condition were compared on the same cardiac biopsy. Eventually, we did not evaluate chlordecone metabolites, such as chlordecol, and sulfonide and glucuronide compounds, which may represent to main active circulating molecule in human toxicity. That chlordecone permeates cardiomyocytes to exert its mitochondrial
<figure><img src="image_12.png" /><figcaption>**Fig. 6.** Proposed mechanical cascade from chlordecone exposure to mitochondrial dysfunction and potential consequences on cardiac function.</figcaption></figure>

uncoupling effect may not be the same in human. It is, indeed, possible that cardiac effects of chlordecone in human may be indirectly related to mechanisms such as increased levels of thyroid and sex-steroid hormones, which have well-described cardiac and mitochondrial action (Ayhan et al., 2021).

## 6. Conclusion

In conclusion, our study has identified mitochondria of human heart as a potential target for the chlordecone toxicity. Owing to the role of mitochondrial dysfunction in many heart diseases, impaired cellular respiration and mitochondrial signaling should be further studied in experimental and clinical settings of human chlordecone poisoning.

Funding.

This research received no external funding.

Institutional Review Board Statement

The study was conducted in accordance with the amended Declaration of Helsinki (https://www.wma.net/policies-post/wma-declaration-of-helsinki/). All patients gave their informed consent for the processing of personal data and tissue collection for scientific research purposes. The study was approved by the French medical ethics committee (CPP: 19-MART-01).

Informed Consent Statement.

Written informed consent was obtained from all subjects involved in the study.

Data Availability Statement.

Research data are available on request at remi.neviere@chumartinique.fr.

### CRediT authorship contribution statement

**Alexia FUNDERE:** Formal analysis, Data curation. **Marie-Daniela DUBOIS:** Formal analysis, Data curation. **Magalie VATIN INAMO:** Formal analysis, Data curation. **Fatima RADOUANI:** Formal analysis, Data curation. **Prisca JALTA:** Formal analysis, Data curation. **Dabor RESIERE:** Conceptualization. **Remi NEVIERE:** Writing – original draft, Methodology, Formal analysis, Conceptualization.

### Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

### References

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