Deficiency of heme oxygenase 1a causes detrimental effects on cardiac function.

Humans lacking heme oxygenase 1 (HMOX1) display growth retardation, haemolytic anaemia, and vulnerability to stress; however, cardiac function remains unclear. We aimed to explore the cardiac function of zebrafish lacking hmox1a at baseline and in response to stress. We generated zebrafish hmox1a mutants using CRISPR/Cas9 genome editing technology. Deletion of hmox1a increases cardiac output and further induces hypertrophy in adults. Adults lacking hmox1a develop myocardial interstitial fibrosis, restrain cardiomyocyte proliferation and downregulate renal haemoglobin and cardiac antioxidative genes. Larvae lacking hmox1a fail to respond to hypoxia, whereas adults are insensitive to isoproterenol stimulation in the heart, suggesting that hmox1a is necessary for cardiac response to stress. Haplodeficiency of hmox1a stimulates non-mitochondrial respiration and cardiac cell proliferation, increases cardiac output in larvae in response to hypoxia, and deteriorates cardiac function and structure in adults upon isoproterenol treatment. Intriguingly, haplodeficiency of hmox1a upregulates cardiac hmox1a and hmox1b in response to isoproterenol. Collectively, deletion of hmox1a results in cardiac remodelling and abrogates cardiac response to hypoxia and isoproterenol. Haplodeficiency of hmox1a aggravates cardiac response to the stress, which could be associated with the upregulation of hmox1a and hmox1b. Our data suggests that HMOX1 homeostasis is essential for maintaining cardiac function and promoting cardioprotective effects.

Moderate hyperuricaemia ameliorated kidney damage in a low-renin model of experimental renal insufficiency.

Uric acid has promoted renal fibrosis and inflammation in experimental studies, but some studies have shown nephroprotective effects due to alleviated oxidative stress. We studied the influence of experimental hyperuricaemia in surgically 5/6 nephrectomized rats. Three weeks after subtotal nephrectomy or sham operation, the rats were allocated to control diet or 2.0% oxonic acid (uricase inhibitor) diet for 9 weeks. Then blood, urine and tissue samples were taken, and renal morphology and oxidative stress were examined. Inflammation and fibrosis were evaluated using immunohistochemistry and real-time PCR (RT-PCR). Remnant kidney rats ingesting normal or oxonic acid diet presented with ~60% reduction of creatinine clearance and suppressed plasma renin activity. Oxonic acid diet increased plasma uric acid levels by >80 µmol/L. In remnant kidney rats, moderate hyperuricaemia decreased glomerulosclerosis, tubulointerstitial damage and kidney mast cell count, without influencing the fibrosis marker collagen I messenger RNA (mRNA) content. In both sham-operated and 5/6 nephrectomized rats, the mast cell product 11-epi-prostaglandin-F2α excretion to the urine and kidney tissue cyclooxygenase-2 (COX-2) levels were decreased. To conclude, hyperuricaemic remnant kidney rats displayed improved kidney morphology and reduced markers of oxidative stress and inflammation. Thus, moderately elevated plasma uric acid had beneficial effects on the kidney in this low-renin model of experimental renal insufficiency.

Tankyrase Inhibition Attenuates Cardiac Dilatation and Dysfunction in Ischemic Heart Failure.

Hyperactive poly(ADP-ribose) polymerases (PARP) promote ischemic heart failure (IHF) after myocardial infarction (MI). However, the role of tankyrases (TNKSs), members of the PARP family, in pathogenesis of IHF remains unknown. We investigated the expression and activation of TNKSs in myocardium of IHF patients and MI rats. We explored the cardioprotective effect of TNKS inhibition in an isoproterenol-induced zebrafish HF model. In IHF patients, we observed elevated TNKS2 and DICER and concomitant upregulation of miR-34a-5p and miR-21-5p in non-infarcted myocardium. In a rat MI model, we found augmented TNKS2 and DICER in the border and infarct areas at the early stage of post-MI. We also observed consistently increased TNKS1 in the border and infarct areas and destabilized AXIN in the infarct area from 4 weeks onward, which in turn triggered Wnt/ß-catenin signaling. In an isoproterenol-induced HF zebrafish model, inhibition of TNKS activity with XAV939, a TNKSs-specific inhibitor, protected against ventricular dilatation and cardiac dysfunction and abrogated overactivation of Wnt/ß-catenin signaling and dysregulation of miR-34a-5p induced by isoproterenol. Our study unravels a potential role of TNKSs in the pathogenesis of IHF by regulating Wnt/ß-catenin signaling and possibly modulating miRNAs and highlights the pharmacotherapeutic potential of TNKS inhibition for prevention of IHF.

Alterations of Cardiac Protein Kinases in Cyclic Nucleotide-Dependent Signaling Pathways in Human Ischemic Heart Failure.

Objectives: Impaired protein kinase signaling is a hallmark of ischemic heart disease (IHD). Inadequate understanding of the pathological mechanisms limits the development of therapeutic approaches. We aimed to identify the key cardiac kinases and signaling pathways in patients with IHD with an effort to discover potential therapeutic strategies. Methods: Cardiac kinase activity in IHD left ventricle (LV) and the related signaling pathways were investigated by kinomics, transcriptomics, proteomics, and integrated multi-omics approach. Results: Protein kinase A (PKA) and protein kinase G (PKG) ranked on top in the activity shift among the cardiac kinases. In the IHD LVs, PKA activity decreased markedly compared with that of controls (62% reduction, p = 0.0034), whereas PKG activity remained stable, although the amount of PKG protein increased remarkably (65%, p = 0.003). mRNA levels of adenylate cyclases (ADCY 1, 3, 5, 9) and cAMP-hydrolysing phosphodiesterases (PDE4A, PDE4D) decreased significantly, although no statistically significant alterations were observed in that of PKGs (PRKG1 and PRKG2) and guanylate cyclases (GUCYs). The gene expression of natriuretic peptide CNP decreased remarkably, whereas those of BNP, ANP, and neprilysin increased significantly in the IHD LVs. Proteomics analysis revealed a significant reduction in protein levels of "Energy metabolism" and "Muscle contraction" in the patients. Multi-omics integration highlighted intracellular signaling by second messengers as the top enriched Reactome pathway. Conclusion: The deficiency in cAMP/PKA signaling pathway is strongly implicated in the pathogenesis of IHD. Natriuretic peptide CNP could be a potential therapeutic target for the modulation of cGMP/PKG signaling.

MiR-185-5p regulates the development of myocardial fibrosis.

BACKGROUND: Cardiac fibrosis stiffens the ventricular wall, predisposes to cardiac arrhythmias and contributes to the development of heart failure. In the present study, our aim was to identify novel miRNAs that regulate the development of cardiac fibrosis and could serve as potential therapeutic targets for myocardial fibrosis. METHODS AND RESULTS: Analysis for cardiac samples from sudden cardiac death victims with extensive myocardial fibrosis as the primary cause of death identified dysregulation of miR-185-5p. Analysis of resident cardiac cells from mice subjected to experimental cardiac fibrosis model showed induction of miR-185-5p expression specifically in cardiac fibroblasts. In vitro, augmenting miR-185-5p induced collagen production and profibrotic activation in cardiac fibroblasts, whereas inhibition of miR-185-5p attenuated collagen production. In vivo, targeting miR-185-5p in mice abolished pressure overload induced cardiac interstitial fibrosis. Mechanistically, miR-185-5p targets apelin receptor and inhibits the anti-fibrotic effects of apelin. Finally, analysis of left ventricular tissue from patients with severe cardiomyopathy showed an increase in miR-185-5p expression together with pro-fibrotic TGF-ß1 and collagen I. CONCLUSIONS: Our data show that miR-185-5p targets apelin receptor and promotes myocardial fibrosis.

Zebrafish Heart Failure Models.

Heart failure causes significant morbidity and mortality worldwide. The understanding of heart failure pathomechanisms and options for treatment remain incomplete. Zebrafish has proven useful for modeling human heart diseases due to similarity of zebrafish and mammalian hearts, fast easily tractable development, and readily available genetic methods. Embryonic cardiac development is rapid and cardiac function is easy to observe and quantify. Reverse genetics, by using morpholinos and CRISPR-Cas9 to modulate gene function, make zebrafish a primary animal model for in vivo studies of candidate genes. Zebrafish are able to effectively regenerate their hearts following injury. However, less attention has been given to using zebrafish models to increase understanding of heart failure and cardiac remodeling, including cardiac hypertrophy and hyperplasia. Here we discuss using zebrafish to study heart failure and cardiac remodeling, and review zebrafish genetic, drug-induced and other heart failure models, discussing the advantages and weaknesses of using zebrafish to model human heart disease. Using zebrafish models will lead to insights on the pathomechanisms of heart failure, with the aim to ultimately provide novel therapies for the prevention and treatment of heart failure.

Association of miR-21-5p, miR-122-5p, and miR-320a-3p with 90-Day Mortality in Cardiogenic Shock.

Cardiogenic shock (CS) is a life-threatening emergency. New biomarkers are needed in order to detect patients at greater risk of adverse outcome. Our aim was to assess the characteristics of miR-21-5p, miR-122-5p, and miR-320a-3p in CS and evaluate the value of their expression levels in risk prediction. Circulating levels of miR-21-5p, miR-122-5p, and miR-320a-3p were measured from serial plasma samples of 179 patients during the first 5-10 days after detection of CS, derived from the CardShock study. Acute coronary syndrome was the most common cause (80%) of CS. Baseline (0 h) levels of miR-21-5p, miR-122-5p, and miR-320a-3p were all significantly elevated in nonsurvivors compared to survivors (p < 0.05 for all). Above median levels at 0h of each miRNA were each significantly associated with higher lactate and alanine aminotransferase levels and decreased glomerular filtration rates. After adjusting the multivariate regression analysis with established CS risk factors, miR-21-5p and miR-320a-3p levels above median at 0 h were independently associated with 90-day all-cause mortality (adjusted hazard ratio 1.8 (95% confidence interval 1.1-3.0), p = 0.018; adjusted hazard ratio 1.9 (95% confidence interval 1.2-3.2), p = 0.009, respectively). In conclusion, circulating plasma levels of miR-21-5p, miR-122-5p, and miR-320a-3p at baseline were all elevated in nonsurvivors of CS and associated with markers of hypoperfusion. Above median levels of miR-21-5p and miR-320a-3p at baseline appear to independently predict 90-day all-cause mortality. This indicates the potential of miRNAs as biomarkers for risk assessment in cardiogenic shock.

Functional imaging with 11C-metomidate PET for subtype diagnosis in primary aldosteronism.

OBJECTIVE: Endocrine Society guidelines recommend adrenal venous sampling (AVS) in primary aldosteronism (PA) if adrenalectomy is considered. We tested whether functional imaging of adrenal cortex with 11C-metomidate (11C-MTO) could offer a noninvasive alternative to AVS in the subtype classification of PA. DESIGN: We prospectively recruited 58 patients with confirmed PA who were eligible for adrenal surgery. METHODS: Subjects underwent AVS and 11C-MTO-PET without dexamethasone pretreatment in random order. The lateralization of 11C-MTO-PET and adrenal CT were compared with AVS in all subjects and in a prespecified adrenalectomy subgroup in which the diagnosis was confirmed with immunohistochemical staining for CYP11B2. RESULTS: In the whole study population, the concordance of AVS and 11C-MTO-PET was 51% and did not differ from that of AVS and adrenal CT (53%). The concordance of AVS and 11C-MTO-PET was 55% in unilateral and 44% in bilateral PA. In receiver operating characteristics analysis, the maximum standardized uptake value ratio of 1.16 in 11C-MTO-PET had an AUC of 0.507 (P = n.s.) to predict allocation to adrenalectomy or medical therapy with sensitivity of 55% and specificity of 44%. In the prespecified adrenalectomy subgroup, AVS and 11C-MTO-PET were concordant in 10 of 19 subjects with CYP11B2-positive adenoma and in 6 of 10 with CYP11B2-positivity without an adenoma. CONCLUSIONS: The concordance of 11C-MTO-PET with AVS was clinically suboptimal, and did not outperform adrenal CT. In a subgroup with CYP11B2-positive adenoma, 11C-MTO-PET identified 53% of cases. 11C-MTO-PET appeared to be inferior to AVS for subtype classification of PA.

Pregnancy-Related Complications in Patients With Fibromuscular Dysplasia: A Report From the European/International Fibromuscular Dysplasia Registry.

Current literature suggests a higher risk of pregnancy-related complications in patients with renal fibromuscular dysplasia (FMD). The aim of our study was to assess the nature and prevalence of pregnancy-related complications in patients subsequently diagnosed with FMD. A call for participation was sent to centers contributing to the European/International FMD Registry. Patients with at least 1 pregnancy were included. Data on pregnancy were collected through medical files and FMD characteristics through the European/International FMD Registry. Data from 534 pregnancies were obtained in 237 patients. Despite the fact that, in 96% of cases, FMD was not diagnosed before pregnancy, 40% of women (n=93) experienced pregnancy-related complications, mostly gestational hypertension (25%) and preterm birth (20%), while preeclampsia was reported in only 7.5%. Only 1 patient experienced arterial dissection and another patient an aneurysm rupture. When compared with patients without pregnancy-related complications, patients with complicated pregnancies were younger at FMD diagnosis (43 versus 51 years old; P<0.001) and had a lower prevalence of cerebrovascular FMD (30% versus 52%; P=0.003) but underwent more often renal revascularization (63% versus 40%, P<0.001). In conclusion, the prevalence of pregnancy-related complications such as gestational hypertension and preterm birth was high in patients with FMD, probably related to the severity of renal FMD. However, the prevalence of preeclampsia and arterial complications was low/moderate. These findings emphasize the need to screen hypertensive women for FMD to ensure revascularization before pregnancy if indicated and appropriate follow-up during pregnancy, without discouraging patients with FMD from considering pregnancy.

Unfavorable Reduction in the Ratio of Endothelin B to A Receptors in Experimental 5/6 Nephrectomy and Adenine Models of Chronic Renal Insufficiency.

Chronic renal insufficiency (CRI) is characterized by increased endothelin 1 (ET-1) synthesis. We studied rat kidney endothelin receptor A (ETA) and receptor B (ETB) expressions after 12 and 27 weeks of 5/6 nephrectomy, and after 12 weeks of 0.3% adenine diet, representing proteinuric and interstitial inflammation models of CRI, respectively. Uric acid and calcium-phosphate metabolism were modulated after 5/6 nephrectomy, while ETA blocker and calcimimetic were given with adenine. Endothelin receptor mRNA levels were measured using RT-qPCR and protein levels using autoradiography (5/6 nephrectomy) or ELISA (adenine model). Both 12 and 27 weeks after 5/6 nephrectomy, kidney cortex ETA protein was increased by ~60% without changes in ETB protein, and the ETB:ETA ratio was reduced. However, the ETB:ETA mRNA ratio did not change. In the adenine model, kidney ETA protein was reduced by ~70%, while ETB protein was suppressed by ~95%, and the ETB:ETA ratio was reduced by ~85%, both at the protein and mRNA levels. The additional interventions did not influence the observed reductions in the ETB:ETA ratio. To conclude, unfavorable reduction in the ETB:ETA protein ratio was observed in two different models of CRI. Therefore, ETA blockade may be beneficial in a range of diseases that cause impaired kidney function.

Vezf1 regulates cardiac structure and contractile function.

BACKGROUND: Vascular endothelial zinc finger 1 (Vezf1) is a transcription factor previously shown to regulate vasculogenesis and angiogenesis. We aimed to investigate the role of Vezf1 in the postnatal heart. METHODS: The role of Vezf1 in regulating cardiac growth and contractile function was studied in zebrafish and in primary cardiomyocytes. FINDINGS: We find that expression of Vezf1 is decreased in diseased human myocardium and mouse hearts. Our experimental data shows that knockdown of zebrafish Vezf1 reduces cardiac growth and results in impaired ventricular contractile response to ß-adrenergic stimuli. However, Vezf1 knockdown is not associated with dysregulation of cardiomyocyte Ca2+ transient kinetics. Gene ontology enrichment analysis indicates that Vezf1 regulates cardiac muscle contraction and dilated cardiomyopathy related genes and we identify cardiomyocyte Myh7/ß-MHC as key target for Vezf1. We further identify a key role for an MCAT binding site in the Myh7 promoter regulating the response to Vezf1 knockdown and show that TEAD-1 is a binding partner of Vezf1. INTERPRETATION: We demonstrate a role for Vezf1 in regulation of compensatory cardiac growth and cardiomyocyte contractile function, which may be relevant in human cardiac disease.

Inhibition of let-7c Regulates Cardiac Regeneration after Cryoinjury in Adult Zebrafish.

The let-7c family of micro-RNAs (miRNAs) is expressed during embryonic development and plays an important role in cell differentiation. We have investigated the role of let-7c in heart regeneration after injury in adult zebrafish. let-7c antagomir or scramble injections were given at one day after cryoinjury (1 dpi). Tissue samples were collected at 7 dpi, 14 dpi and 28 dpi and cardiac function was assessed before cryoinjury, 1 dpi, 7 dpi, 14 dpi and 28 dpi. Inhibition of let-7c increased the rate of fibrinolysis, increased the number of proliferating cell nuclear antigen (PCNA) positive cardiomyocytes at 7 dpi and increased the expression of the epicardial marker raldh2 at 7 dpi. Additionally, cardiac function measured with echocardiography recovered slightly more rapidly after inhibition of let-7c. These results reveal a beneficial role of let-7c inhibition in adult zebrafish heart regeneration.

Circulating levels of microRNA 423-5p are associated with 90 day mortality in cardiogenic shock.

AIMS: The role of microRNAs has not been studied in cardiogenic shock. We examined the potential role of miR-423-5p level to predict mortality and associations of miR-423-5p with prognostic markers in cardiogenic shock. METHODS AND RESULTS: We conducted a prospective multinational observational study enrolling consecutive cardiogenic shock patients. Blood samples were available for 179 patients at baseline to determine levels of miR-423-5p and other biomarkers. Patients were treated according to local practice. Main outcome was 90 day all-cause mortality. Median miR-423-5p level was significantly higher in 90 day non-survivors [median 0.008 arbitrary units (AU) (interquartile range 0.003-0.017) vs. 0.004 AU (0.002-0.009), P = 0.003]. miR-423-5p level above median was associated with higher lactate (median 3.7 vs. 2.4 mmol/L, P = 0.001) and alanine aminotransferase levels (median 68 vs. 35 IU/L, P < 0.001) as well as lower cardiac index (1.8 vs. 2.4, P = 0.04) and estimated glomerular filtration rate (56 vs. 70 mL/min/1.73 m2 , P = 0.002). In Cox regression analysis, miR-423-5p level above median was associated with 90 day all-cause mortality independently of established risk factors of cardiogenic shock [adjusted hazard ratio 1.9 (95% confidence interval 1.2-3.2), P = 0.01]. CONCLUSIONS: In cardiogenic shock patients, above median level of miR-423-5p at baseline is associated with markers of hypoperfusion and seems to independently predict 90 day all-cause mortality.

Pooled analysis of Phase III trials indicate contrasting influences of renal function on blood pressure, body weight, and HbA1c reductions with empagliflozin.

Sodium glucose cotransporter 2 (SGLT2) inhibitors reduce HbA1c, blood pressure, and weight in patients with type 2 diabetes. To investigate the effect of renal function on reductions in these parameters with the SGLT2 inhibitor empagliflozin, we assessed subgroups by baseline estimated glomerular filtration rate (eGFR; Modification of Diet in Renal Disease) in pooled data from five 24-week trials of 2286 patients with type 2 diabetes randomized to empagliflozin or placebo. Reductions in HbA1c with empagliflozin versus placebo significantly diminished with decreasing baseline eGFR. Reductions in systolic blood pressure (SBP) with empagliflozin were maintained in patients with lower eGFR. The mean placebo-corrected changes from baseline in systolic blood pressure at week 24 with empagliflozin were -3.2 (95% confidence interval -4.9,-1.5) mmHg, -4.0 (-5.4, -2.6) mmHg, -5.5 (-7.6, -3.4) mmHg, and -6.6 (-11.4, -1.8) mmHg in patients with an eGFR of 90 or more, 60 to 89, 30 to 59, and under 30 ml/min/1.73m2, respectively. Similar trends were observed for diastolic blood pressure. Weight loss with empagliflozin versus placebo tended to be attenuated in patients with a lower eGFR. Results were consistent in a 12-week ambulatory blood pressure monitoring trial in 823 patients with type 2 diabetes and hypertension. Thus, unlike HbA1c reductions, systolic blood pressure and weight reductions with empagliflozin are generally preserved in patients with chronic kidney disease.

Carbon monoxide releasing molecule improves structural and functional cardiac recovery after myocardial injury.

Carbon monoxide (CO), produced by heme oxygenase-1 (HO-1), is an endogenous paracrine factor involved in the regulation of cardiovascular structure and function. We studied the effects of a synthetic CO releasing molecule (CORM-3) on cardiac recovery and myocardial microRNA expression after myocardial infarction (MI). Male Wistar rats with MI (n = 75) or sham-operated controls (n = 75) were treated from day 4 to day 14 after MI either with synthetic CORM-3 or with inactive iCORM and killed 2, 4 or 8 weeks post-MI. Infarct size, vascular and capillary densities, the amount of cardiomyocytes in the infarct area, and cardiomyocyte proliferation and apoptosis were determined. PCR was used for microRNA and mRNA quantification, western blotting to evaluate protein expression and echocardiography to assess cardiac structure and function. CORM-3 treatment increased vascular density (P< 0.05 vs. iCORM) and the proportion of cardiomyocytes (P< 0.05 vs. iCORM) in the infarct area. Ejection fraction improved (P< 0.05) and left ventricular volumes decreased (P< 0.05) in CORM-3 treated MI groups compared to iCORM treatment. CORM-3 treatment decreased the amount of proliferating Ki67 positive cardiomyocytes in the infarct/border area at week 2 after MI compared to iCORM treatment, whereas the amount of apoptotic cardiomyocytes did not differ between CORM-3 and iCORM groups. Compared to iCORM treatment, CORM-3 decreased expression on miR-206 in the remote area at week 2 after MI. The CO releasing molecule CORM-3 improved structural and functional cardiac recovery after MI. Modulation of HO-1-CO axis may prove novel drug targets to facilitate cardiac recovery after myocardial injury.

Blood pressure response to renal denervation is correlated with baseline blood pressure variability: a patient-level meta-analysis.

BACKGROUND: Sympathetic tone is one of the main determinants of blood pressure (BP) variability and treatment-resistant hypertension. The aim of our study was to assess changes in BP variability after renal denervation (RDN). In addition, on an exploratory basis, we investigated whether baseline BP variability predicted the BP changes after RDN. METHODS: We analyzed 24-h BP recordings obtained at baseline and 6 months after RDN in 167 treatment-resistant hypertension patients (40% women; age, 56.7 years; mean 24-h BP, 152/90 mmHg) recruited at 11 expert centers. BP variability was assessed by weighted SD [SD over time weighted for the time interval between consecutive readings (SDiw)], average real variability (ARV), coefficient of variation, and variability independent of the mean (VIM). RESULTS: Mean office and 24-h BP fell by 15.4/6.6 and 5.5/3.7 mmHg, respectively (P < 0.001). In multivariable-adjusted analyses, systolic/diastolic SDiw and VIM for 24-h SBP/DBP decreased by 1.18/0.63 mmHg (P ≤ 0.01) and 0.86/0.42 mmHg (P ≤ 0.05), respectively, whereas no significant changes in ARV or coefficient of variation occurred. Furthermore, baseline SDiw (P = 0.0006), ARV (P = 0.01), and VIM (P = 0.04) predicted the decrease in 24-h DBP but not 24-h SBP after RDN. CONCLUSION: RDN was associated with a decrease in BP variability independent of the BP level, suggesting that responders may derive benefits from the reduction in BP variability as well. Furthermore, baseline DBP variability estimates significantly correlated with mean DBP decrease after RDN. If confirmed in younger patients with less arterial damage, in the absence of the confounding effect of drugs and drug adherence, baseline BP variability may prove a good predictor of BP response to RDN.

Fetal Microsatellite in the Heme Oxygenase 1 Promoter Is Associated With Severe and Early-Onset Preeclampsia.

Preeclampsia is a vascular pregnancy disorder that often involves impaired placental development. HO-1 (heme oxygenase 1, encoded by HMOX1) is a stress response enzyme crucial for endothelial and placental function. Long version of the guanine-thymine (GTn) microsatellite in the HMOX1 promoter decreases HO-1 expression, and the long maternal repeat is associated with late-onset preeclampsia. Our aim was to study whether the length of fetal repeat is associated with mother's preeclampsia, whether the length of fetal and maternal repeats affect HO-1 levels in placenta and maternal serum, and whether HO-1 levels are altered in preeclampsia. We genotyped the repeat in the cord blood of 609 preeclamptic and 745 nonpreeclamptic neonates. HO-1 levels were measured in 36 placental samples, and in the first (222 cases/243 controls) and third (176 cases/53 controls) pregnancy trimester serum samples using enzyme-linked immunosorbent assay. The long fetal GTn repeat was associated with preeclampsia and its severe and early-onset subtypes. Interaction analysis suggested the maternal and fetal effects to be independent. Placental or serum HO-1 levels were not altered in preeclamptics, possibly reflecting heterogeneity of preeclampsia. Carriers of the long fetal and maternal repeats had lower placental and serum HO-1 levels, respectively, providing functional evidence for the association. We conclude that the long fetal GTn repeat may increase mother's risk for especially severe and early-onset preeclampsia. The fetal and maternal risk alleles likely predispose to different disease subtypes.