Sex Differences in Pulmonary (Arterial) Hypertension – Does it Matter?

Abstract

Purpose of review:

This review synthesizes the current prevailing theories behind the “sex paradox” or “sex puzzle” in pulmonary arterial hypertension (PAH), a disease marked by sexual dimorphism. To a lesser extent, we also review sex differences in other forms of pulmonary hypertension (PH).

Recent Findings:

While more females than males develop PAH worldwide, female sex is associated with improved right ventricular (RV) function and survival. We review the role of sex chromosomes and sex hormones and their relationships to genomic and epigenetic regulation, immune function, and RV function, sex-based differences in therapeutic response and social determinants of health and intersectionality with gender in PAH pathobiology, prevalence and outcomes. We include experimental studies and observational human data that have led to the study of sex hormone modulation as a treatment strategy in PAH, with recently completed clinical trials. In addition, we explore potential future directions to help understand the mechanisms that underpin sex-biases in pulmonary vascular disease, as well as those that may inform potential therapeutic targets.

Summary:

Survival in PAH depends on RV function, and females have improved survival despite increased prevalence for reasons that remain unclear. While knowledge gaps remain, recent advancements offer promise and many future directions.

Introduction

Pulmonary arterial hypertension (PAH) has diverse etiologies, yet female sex is a unifying clinical risk factor in many subtypes and PAH is sexually dimorphic in terms of penetrance, severity, right ventricular (RV) function, response to therapy, and survival. Modern registries have demonstrated that as many as 80% of PAH patients are women and that sex bias may change with age [1, 2*, 3]. Recent studies have confirmed that female sex is independently associated with a lower risk of long-term mortality [4*,5**]. Decades of research has sought to understand the mechanisms that contribute to these observations. While less well studied, other forms of pulmonary hypertension (PH) including PH due to left heart, chronic lung diseases and possibly chronic thromboembolic PH, also have evidence of sexual dimorphism.

The “estrogen paradox,” was coined in 2010 by Tofovic [6]. Now, the terms “sex paradox” or “sex puzzle” are used, acknowledging that prevailing theories to explain why more women develop disease yet have improved survival go well beyond estrogen alone and include additional sex hormones and their metabolites, sex chromosome contributions, and RV adaptive responses, among others. Here, we provide a concise review of sex and gender differences in PAH (see Figure 1) (and the more limited data in PH), progress in understanding these differences, and gaps that remain. For the purposes of this review, we will use the terms “female” and “male” when referring to biological sex at birth, “women” and men” when referring to gender, and acknowledge that these are not interchangeable nor are they binary.

(Original) Figure 1 – Sex Differences in Pulmonary Arterial Hypertension.

A schematic of the major sexually dimorphic features in pulmonary arterial hypertension (PAH) beginning with murine models of PH. SRY = Sex-determining region Y-Chromosome, also called testis determining factor (TDF), a transcription factor encoded by the SRY/Sry gene on the Y-Chromosome (Yp or short arm). UTY = ubiquitously transcribed tetratricopeptide repeat containing Y-linked, a histone demethylase enzyme from the UTY/Uty gene on the Y-chromosome (Yq or long arm). Uty has been implicated in protection against PH in murine studies [18]; XCI = X-chromosome inactivation; Xist = long non-coding RNA called X-inactive specific transcript. Once Xist is activated, XCI occurs through DNA methylation, histone modification, and chromatin compaction to form Barr bodies [7]. BMPR2 = Bone morphogenetic protein receptor type II, a gene that encodes for BMPR2, a serine/threonine receptor kinase, and has sexually dimorphic features and binds members of the TGFβ superfamily of ligands [28,104]. T = testosterone; E2 = 17β-estradiol; DHEA = dehydroepiandrosterone and DHEA-S dehydroepiandrosterone sulfate are hormones made primarily in the adrenal gland and are precursors for androgens and estrogens; 16α-OHE1 is 16-alpha hydroxyestrone, a highly estrogenic metabolite of estradiol (E2) and estrone (E1), and can also be formulated via adipocytes production of estrone. Aromatase – the enzyme that converts androgens to estrogens; Cortisol – the precursor steroid of sex hormones [29,30]. Lifecycle events and changes – in females, these are namely the menstrual cycle with hormonal fluctuations, hormonal contraceptives, pregnancy, and the transition to menopause. In females, the aging sex hormone profile is marked by increase in gonadotropin releasing hormones (high LH, high FSH), with low E2, low progesterone (P4), and low anti-Müllerian hormone (AMH). There are also less dramatic but consistent decreases in DHEA-S. Testosterone is largely stable in the female. Menopause is marked by large hormonal fluctuations. Aging in males is also marked by hormonal changes with reduction in testosterone and DHEA-S. Created in BioRender.

Sex chromosomes and genomic sex

Biological discussions around sex and sexually dimorphic conditions begin with the X- and Y-chromosomes. Excluding the mature erythrocyte, every cell in the human body has sex chromosomes. Genomic sex impacts micro- and macro-environments across both healthy and disease states, but to-date this has been understudied in pulmonary vascular disease.

Sex chromosomes comprise approximately five percent of the total human genome. In the karyotypical 46,XX female, two X-chromosomes result in a “double dose” of X-linked genes, which is mitigated by a master epigenetic regulator X-inactive specific transcript (Xist), a long-non-coding RNA. Xist regulates X-linked gene expression via X-chromosome inactivation, silencing one of the X-chromosomes. X-chromosome inactivation is crucial for normal female development [7], but approximately 15-25% of X-linked genes escape Xist and evade silencing [8,9]. Called “escapees,” these genes have been linked to autoimmune and sex-biased diseases that are also female predominant and relevant to PAH [10*, 11, 12*, 13*].

Studies to-date of the role of Xist in PAH female predominance are limited. In a study by Qin et al, pulmonary artery endothelial cell (PAEC) Xist expression was higher in PAECs from female PAH subjects compared with female control PAECs, male PAH and control PAECs. ELK1, an X-linked gene implicated in plexiform arteriopathy, expression was upregulated in female PAH-PAECs compared with healthy controls and all males [14]. The same group showed that a subset of male PAH patients had increased Xist expression in PAECs from explanted lungs [15*]. A recently published review examines sex differences in PAH and other human diseases through the lens of Xist [16**].

Once thought to be involved only in male phenotypic expression, the Y-chromosome is far more complex, with about 100 protein-coding genes, including SRY and UTY. Li et al was the first group to examine the role of sex chromosomes in PAH and discover that the Y-chromosome is protective in hypoxia-induced PH. They leveraged the four core genotypes mouse model, which allows for the transgenic manipulation of sex gonads, sex chromosomes and their dose [17]. In a subsequent study, knockdown of Uty resulted in the expression of proinflammatory chemokines Cxcl9 and Cxcl10, which triggered endothelial cell death and more severe PH [18]. In human PAH fibroblasts, SRY has been shown to bind to and regulate bone morphogenetic protein receptor type II (BMPR2) expression in a dose dependent manner [19]. While this preclinical evidence provides some support for the epidemiologic observation that men are less likely to develop PAH, translational human studies in this area remain lacking.

While difficult to study in a rare disease like PAH, loss of the Y-chromosome in blood has been linked to an increased risk of major cardiovascular events in males [20]. In two large cohort studies, the presence of any supernumerary sex chromosome aneuploidy was associated with a small but statistically significant increase in the risk of venous thromboembolism [21]. Klinefelter syndrome (47,XXY) individuals have increased risk of systemic lupus erythematosus (SLE), systemic sclerosis (SSc), Sjögrens and idiopathic inflammatory myopathies, autoimmune diseases traditionally associated with female sex [22,23] and PAH [24*]. These observations may be linked to a “double dose” of the X-chromosome. Rigorously designed studies to address this gap in patients with pulmonary vascular disease are needed.

Sex differences and major causal pathways in PAH

All major pathobiologic pathways in PAH intersect with sex hormone signaling. This includes but is not limited to endothelin-1 (ET-1), nitric oxide, cyclic guanosine monophosphate, as well as prostaglandin I₂, activin, and TGF-beta signaling. Given the limited scope of this review, we will focus on BMPR2’s key role in sex differences in PAH [25].

In heritable PAH (HPAH) due to BMPR2 mutations, penetrance is incomplete and varies by sex. Female carriers of BMPR2 mutations are more likely to develop PAH than male carriers; the risk of PAH is ~42% in females and ~14% in males [26]. There is conflicting data on whether sex bias exists in idiopathic and drug and toxin-associated PAH patients with BMPR2 mutations. In a meta-analysis of 1550 patients with HPAH, idiopathic PAH, and anorexigen-associated PAH, BMPR2 mutation rate did not vary by sex; nor did sex influence the relationship between BMPR2 mutation status and mortality [27]. However, in a different meta-analysis of 17 clinical trials among 2198 participants with idiopathic and heritable PAH, BMPR2 mutation rate was higher in males compared with females [28]. These studies suggest that sex chromosomes or the hormonal milieu (or both) may serve as an additional “hit” for individuals who are genetically preconditioned to develop PAH.

There is known crosstalk between BMPR2, estrogen signaling and metabolism, and disease penetrance. Estrogens are largely metabolized by the liver through cytochrome (CYP) 450 enzymes to different metabolites which have biologically distinct and tissue specific properties, discussed thoroughly elsewhere [29,30*]. One such estrogen metabolite, 16-alpha hydroxyestrone (16α-OHE1), is highly estrogenic and generated via 17β-estradiol (E2) conversion to estrone, or via adipocyte production of estrone, the primary estrogen in postmenopausal females (Figure 2). BMPR2 mutant mice treated with 16α-OHE1 increased the risk of PH and disease severity and was associated with metabolic syndrome [31]. Further, CYP1B1 activity was increased in human PAH lungs and PH mice [32]. Genetic variation in CYP1B1 modified the risk of PAH penetrance among BMPR2 mutation carriers; female affected mutation carriers were more likely to have a 16α-OHE1-dominant urinary estrogen metabolite profile [33]. This same group demonstrated that BMPR2 expression is suppressed through direct promoter binding by estrogen receptor alpha (ERα) [34]. A second study confirmed that estrogen metabolism is linked to PAH penetrance, noting that females with BMPR2 mutations who preferentially metabolized estradiol to 16α-OHE1 developed PAH compared with females who preferentially metabolized to 2- or 4-estrogens [33,35,36]. 16α-OHE1 disrupts cellular metabolism through the upregulation of the microRNA-29 (miR-29) cluster, which downregulates several metabolic genes leading to decreased fatty acid oxidation, increased glycolysis, oxidative stress and disrupted mitochondrial function. Furthermore, 16α-OHE1 can covalently bind to estrogen receptors [6,37], potentially causing prolonged estrogenic stimulus. Aging and several lifestyle risk factors such as obesity, smoking, diet and stress influence the balance of estrogen metabolism, and the generation of active estrogen metabolites [38,39] which may in turn influence PAH clinical phenotypes discussed below.

(NOT ORIGINAL) Figure 2 – Sex hormone synthesis and estrogen metabolism –

FROM COMPREHENSIVE PHYSIOLOGY – (Hester, Ventetuolo, Lahm) (52) Figure 3 in original publication. Full citation: Hester J, Ventetuolo C, Lahm T. Sex, Gender, and Sex Hormones in Pulmonary Hypertension and Right Ventricular Failure. Compr Physiol. 2019;10(1):125-170. Published 2019 Dec 18. doi:10.1002/cphy.c190011

Immune dysregulation, autoimmunity, and sex differences in PAH

Autoimmune and connective tissue disease associated PAH is significantly more prevalent in females. At the same time, immune dysregulation, inflammation and autoimmunity play important roles in the susceptibility and pathogenesis of PAH [40,41,42*]. There are several lines of evidence connecting the immune system (and its dysregulation) to sex hormone metabolism and signaling as well as cellular karyotype which may be relevant in pulmonary vascular disease.

Pro-inflammatory cytokines strongly induce aromatase (the major enzyme responsible for androgen to estrogen conversion), and peripheral E2 synthesis [29,43,44]. Estrogen receptors (ER) are expressed on lymphoid tissue, in lymphocytes, macrophages and dendritic cells. ERα is highly expressed in T cells while ERβ is upregulated in B cells [45,46]. E2 affects many aspects of innate immunity, and ERα signaling promotes T cell activation and proliferation and contributes to T cell-mediated autoimmune inflammation [29,47]. In experimental PH, female rats lacking normal regulatory T cell (Treg) activity exhibited greater inflammation, more dramatic RV microvascular dropout and increased periarticular fibrosis compared with male rats with PH; female rats also had decreased systemic levels of prostacyclin, changes which could be prevented with Treg immune reconstitution [48].

Finally, many genes on the X-chromosome regulate immune function. Evidence suggests that because of incomplete X-chromosome inactivation, females have more expressed immune function genes compared with XY individuals, resulting in a more robust immune system potentially more prone to unchecked activation [49-51]. While both immunity, sex steroid signaling and metabolism, and (to a more limited degree) sex chromosomes have been studied independently in pulmonary vascular disease, there remain gaps in our understanding of the cross talk between these pivotal themes and PAH.

Sex hormones, their receptors and contributions to PAH

There is now an extensive preclinical literature on sex hormones, their metabolism and receptors in pulmonary vascular disease, which is beyond the scope of the current review but is discussed in detail here [52]. Briefly, initial studies of specific sex hormones focused on 17β-estradiol or E2, the primary estrogen synthesized in the ovary in females of reproductive age [6,29]. Of the estrogen receptors (ERα, ERβ, and GPR30), ERα and ERβ have been implicated in PH and are known to be expressed in PAECs, pulmonary artery smooth muscle cells (PASMCs), and fibroblasts, as well as other lung cells, and in cardiomyocytes, cardiac endothelial cells, and fibroblasts [52]. ERα has been linked to proliferative and inflammatory effects, particularly in PASMCs, while ERβ may exert an anti-proliferative and vasoprotective role [52-55]. Conversely, recent work by Frump et al demonstrates that loss of ERα results in more severe PH and in human PAH PAECs, ERα abundance is decreased, suggesting impaired ERα signaling in PAH [56,57]. Estradiol synthesis and metabolism are also regulated locally and may be compartment specific. For example, aromatase is abundantly expressed in the smooth muscle layer of pulmonary arteries in healthy subjects and PAH patients [58]. Increased expression of both aromatase and CYP1B1 has been demonstrated in PAH lungs as compared to controls [32,52,58,59].

A number of cohort and case-control studies have characterized circulating hormone profiles in PAH. Higher levels of E2, greater E2/testosterone ratio, and lower levels of dehydroepiandrosterone-sulfate (DHEA-S) was demonstrated in males with PAH as compared to age and body mass index (BMI) matched controls, and higher E2 and lower DHEA-S was associated with more severe PAH metrics [60]. In a follow-up study of men with PAH from China, similar hormone profiles were observed [61]. In studies in postmenopausal females, and premenopausal females over the course of the menstrual cycle, the high E2, low DHEA-S signature in PAH persisted, and lower levels of DHEA-S were associated with worse PAH metrics including RV dysfunction [60,62,63]. During all phases of the menstrual cycle, women with PAH had higher levels of E2 and lower levels of DHEA-S as compared to healthy controls, but the association with E2 and PAH endpoints was variable (associated with better or worse PAH measures, depending on phase of the cycle), while lower levels of DHEA-S were consistently associated with worse outcomes [62]. Lower levels of DHEA-S in PAH have been demonstrated over several studies now, including in unsupervised metabolomics studies, and are associated with poor survival [64*,65,66] (Figure 2).

Clinical trials of hormonal manipulation as a treatment strategy in PAH

Anastrozole, an approved treatment for breast cancer, blocks aromatase, thus preventing androgen conversion to estrogens. Genetic variation in aromatase is associated with the risk of portopulmonary hypertension [67] and anastrozole is protective in animal PH models [58]. These observations, along with studies confirming that PAH patients have higher circulating levels of E2, led to clinical trials of anastrozole in PAH. In a pilot clinical trial in postmenopausal women and men with PAH, treatment with anastrozole for 12 weeks reduced serum E2 levels by 40%, significantly increased six-minute walk distance (6MWD), and had no effect on RV function measured by echocardiography [68]. In the follow-up Phase II multisite randomized clinical trial PHANTOM, postmenopausal women and men with PAH were randomized to receive anastrozole vs. placebo for 12 months. Anastrozole had no effect on 6MWD or other PAH end points compared with placebo [69*]. Potential explanations for these discrepant findings of the two anastrozole trials include the longer duration of PHANTOM, treatment of all subjects with vitamin D as part of the PHANTOM protocol to prevent anastrozole-associated bone loss, differences in the study samples, or a false positive result in the pilot study.

Selective estrogen receptor modulators (SERMs) are a class of compounds that selectively modulate estrogen receptors in a tissue-specific manner, acting as estrogen agonists in some tissues and antagonists in others. In female monocrotaline-PH rats, chronic administration of raloxifene, a SERM, attenuated the progression of PH in intact and ovariectomized female rats, and these rats displayed reduced RV ET-1 expression [70]. A phase II clinical trial of tamoxifen to treat PAH in pre- and post-menopausal women with PAH has recently been completed (NCT03528902) [71]. Women with PAH also have evidence of increased ERα in pulmonary arterioles [55]. Fulvestrant, an ERα inhibitor approved for the treatment of breast cancer, was given in an open-label pilot study to five postmenopausal women with PAH and was associated with a nonsignificant improvement in 6MWD, no change in tricuspid annular plan systolic excursion, but possible improvement in RV stroke volume, despite no changes in lung uptake of ¹⁸F-fluoroestradiol on positron emission tomography-computed tomography imaging [72]. Finally, preclinical models demonstrating DHEA’s beneficial effect on pulmonary vascular and RV function as well as observational human data illustrating DHEA-S deficiency in patients with PAH led to the design and conduct of EDIPHY (Effects of Dehydroepiandrosterone in Pulmonary Hypertension) randomized crossover trial in women and men with PAH (NCT03648385) [73]. The primary end point for this 42-week trial is RV structure and function as assessed by cardiac magnetic resonance imaging; results are anticipated shortly. Table 1 summarizes the clinical trials to-date which have targeted sex hormones or sex hormone signaling.

(Original) Table 1 –

Clinical Trials Targeting Sex Hormones or Signaling

Clinical Trial	Drug	Mechanism	Objective(s)	Subjects	Duration	Status	Outcomes	Dates	Ref
AIPH - (NCT01545336) - Phase II - Randomized, double-blind, placebo-controlled study at two centers	Anastrozole	Aromatase inhibitor - Blocks conversion of androgens to estrogens	Determine the safety of anastrozole in PAH; Percent change in baseline E2 levels; Change from baseline TAPSE	18 PAH patients (men and post-menopausal women) on background therapy were randomized to Anastrozole 1mg or matching placebo in a 2:1 ratio	3 months	Completed	Anastrozole significantly reduced E2 levels compared with placebo but no difference in TAPSE. Increase in 6MWD; Safe, no adverse events	2012-2015	[67]
PHANTOM - (NCT03229499) - Phase II - Randomized, double-blind, placebo-controlled study at 7 centers	Anastrozole	Aromatase inhibitor - Blocks conversion of androgens to estrogens	Improvement in 6MWD; additional secondary aims	84 post-menopausal women and men with PAH on background therapy. 41 randomized to placebo (3 withdrew), 43 randomized to Anastrozole 1mg PO once daily (2 withdrew).	12 months	Completed	No difference in 6MWD at 3, 6, and 12-months compared with baseline; Anastrozole was safe; no adverse effects	2017-2022	[69]
T3PAH - (NCT03528902) - Phase II - Randomized, double-blind, placebo-controlled study at one center	Tamoxifen	Selective Estrogen Receptor Modulator (SERM) that competitively binds to ERs on target tissues	Change from baseline TAPSE; Additional secondary aims	18 subjects enrolled in a 1:1 manner, randomized to tamoxifen 20mg PO daily vs placebo	24 weeks	Completed	TBD	2018-2022	[71]
ERA-PAH - (NCT02911844) - Proof of concept, open-label, single center	Fulvestrant	Estrogen-receptor alpha antagonist -- dose-related down-regulation of ERs	A) Change in 6MWD, plasma NT-proBNP, HPCs, estradiol levels, and other blood biomarkers; change in TTE evaluation of right ventricular function (RV systolic pressure, stroke volume, and TAPSE). B) To evaluate lung uptake of estradiol bindings on PET using separate protocol and consent (NCT02899533).	Five post-menopausal women with PAH (two IPAH, two CTD-PAH, one HIV-PAH) with WHO-FC II and III symptoms. Every subject received Fulvestrant administered in a 500-mg dose intramuscularly on Days 0, 14, 28, and 56	9 weeks	Completed	A) Fulvestrant may be associated with numerically higher 6MWD, increasing stroke volume, and a decrease in 16OHE2 (estriol/E3) and circulating HPCs. No changes in echocardiographic measures, biomarkers of insulin resistance or inflammation. B) No change in 18F-FES uptake in the lungs on PET.	2017-2018	[72]
EDIPHY - (NCT03648385) - Single-center, randomized, cross-over, placebo-controlled	DHEA	DHEA and DHEA-S are adrenally secreted precursor hormones of androgens and estrogens*	Assess if DHEA supplementation improves RV contractility and maladaptive response as measured by cardiac MRI; Change in serum biomarkers	Goal of 26 PAH patients (men, pre and post-menopausal women)	Two 18-week treatment periods with 4-week washout	Completed	Expected shortly	2019 - Current	[73]

Key:

AIPH = Anastrozole in Patients With PAH

PHANTOM = Pulmonary Hypertension and Anastrozole Trial

T3PAH = Tamoxifen Therapy to Treat PAH

ERA-PAH = Estrogen Receptor Antagonist in Patients with PAH

EDIPHY = Effects of DHEA in PH

DHEA = dehydroepiandrosterone

TTE = transthoracic echocardiography

TAPSE = tricuspid annular plane systolic excursion

E2 = 17 beta-estradiol

ER = Estrogen Receptor

NT-proBNP = N-terminal pro-brain natriuretic peptide

HPC = hematopoietic progenitor cells

TBD = To be determined

16OHE2 = 16α-hydroxyestradiol (also known as estriol, or E3)

18F-FES = an established probe for estradiol binding to estrogen receptors

PET-CT = positron emission tomography–computed tomography

6MWD = Six-minute walk distance

WHO-FC = World Health Organization-Functional Class

IPAH = idiopathic pulmonary arterial hypertension

CTD-PAH = connective tissues disease associated pulmonary arterial hypertension

HIV-PAH = human immunodeficiency virus associated pulmonary arterial hypertension

^*DHEA and DHEA-S are secreted primarily from the adrenal gland (zona reticularis), with small contributions from the ovary (theca cells) and testes (Leydig cells)

Sex-based differences in PH and PAH phenotypes

In healthy subjects, the normal mean pulmonary artery pressure (mPAP) at rest does not vary by sex [74]. However, in PAH, men have worse hemodynamics than women (higher mean pulmonary artery pressure (mPAP), right atrial pressure (RAP) and pulmonary vascular resistance (PVR), and lower cardiac index (CI), although some of these differences appear to attenuate after age 45, implicating potential lifecycle changes such as menopause [75]. In a follow-up study of 6,633 clinical trial subjects, baseline differences in hemodynamics were confirmed (females had lower RAP, mPAP and higher CI); age and BMI had differential effect modification that varied by sex on PAH end points including hemodynamics and 6MWD [76]. In the REVEAL registry, men had higher mPAP and higher RAP, and in fact being a male over 60 increases an individual’s REVEAL risk assessment score and is associated with poorer survival [1,77]. In REVEAL, women were more likely to have CTD-PAH, congenital heart disease-PAH, and thyroid disease, while men were more likely to have portopulmonary hypertension and HIV associated PAH [1]. Biological sex and sex hormones have been strongly implicated in portopulmonary hypertension phenotypes [67,78]. Modern PH registries from around the globe and their reported proportion of included females are included in Table 2.

Table 2 (Original) –

List of Recent and Historic Pulmonary Arterial Hypertension Registries

Registry	North America
	Total number patients	Median age (years)		Female %		Subtypes PAH	Dates Represented	Other notes	Ref
		PAH	IPAH	PAH	IPAH
NIH PPH (USA)	187	36	n/a	63%	n/a	HPAH = 12 (5=F, 7=M)	1981-1985	F:M ratio of 1.7:1 which was changed to percent^; 15 out of 16 patients with Raynaud's were female [106]	[105] [106]
PHC - Pulmonary Hypertension Connection (USA)	578	48	n/a	77.0%	n/a	IPAH = 48; CTD = 30; CHD = 11; PoPH = 7; Anorexigens = 3; HIV = 1	1982-2006	n/a	[107]
REVEAL (USA)	2525	53	53	79.5%	80.3%	IPAH = 1166; all APAH = 1280 -- breakdown CTD = 639; CHD = 250; PoPH = 136; Drug/toxin = 134	2006-2007	F:M ratio = 4.07:1 overall; 3.06:1 among newly diagnosed IPAH/HPAH; n=216 pediatric patients (64% F, median age dx 7 years)	[77]
PHAR (USA)	1891	56	n/a	75.0%	76.3%	IPAH = 817 (624=F, 193=M); HPAH = 55 (41=F, 14=M); CTD = 617 (532=F, 85=M); PoPH = 127 (67=F, 60=M); Drug/toxin = 243 (152=F, 91=M); HIV = 32 (9=F, 23=M)	2015 - current	On-going, active registry with new publications	[2]
USPHSR (USA)	499	51.6	49.1	79.0%	74.0%	IPAH = 218 (162=F); HPAH = 22; (18=F); PVOD/PCH = 3 (2=F); APAH = 256 (210=F)	2016-2018	APAH subtypes: CTD = 171 (154=F); CHD = 30 (21=F); PoPH = 22 (9=F) Drugs/toxin = 24 (19=F); HIV = 5 (3=F); Other = 4 (4=F)	[108]
REMEHIP - The Pulmonary Hypertension Mexican Registry (Mexico)	619	43 (CHD = 39; CTD = 49)	43.7	81.9%	85% (IPAH/HPAH)	IPAH/HPAH = 249 (207=F, 85%); CTD = 111 (99=F, 88%); CHD = 249 (189=F, 76%); Other: (anorexigen = 3 (2=F), PoPH = 3 (2=F), and HIV = 3 (2=F) were infrequent (0.5%) and excluded	2015 - 2018	Largest registry in Latin America; CTEPH = 123 (69=F); age 44. Pediatric PAH = 133 (68=F, 51%), mean age 9; IPAH = 38; CHD = 94, CTD = 1	[109]
CPHR - Canadian Pulmonary Hypertension Registry (Canada)	125	61.5	n/a	69.6%	n/a	IPAH = 80; CTD = 33; CHD = 10; PoPH = 2; Drug/toxin = 0; HIV = 0	2009-2015	On-going, active registry	[110]
	267	60.9	n/a	64.7%	n/a	IPAH = 149; CTD = 86; CHD = 14; PoPH = 5; Drug/toxin = 12; HIV = 1	2015-2021
Registry	Europe
	Total number patients	Median age (years)		Female %		Subtypes PAH (if known)	Dates Represented	Other	Ref
		PAH	IPAH	PAH	IPAH
French Registry	2879	61	n/a	60.3%	n/a	IPAH = 1094; HPAH = 137; CTD = 781; SSc = 603; CHD 23; PoPH = 525; HIV = 89; Drug/toxin = 230	Jan 2009 - Dec 2020	Excluded those with missing data (WHO-FC, 6MWD, NT-proBNP/BNP), PVOD, unrepaired CHD, & Eisenmenger syn	[111]
REHAP (Spain)	3169	53.0 (all pts PAH + CTEPH)	50.6 (I/HPAH)	67.8% ^	68% (I/HPAH)	I/HPAH = 940 (640=F, 300=M); CTD = 728 (626=F, 102=M); CHD = 752 (512=F, 240 = M); PoPH = 314 (140=F, 174=M); PVOD = 161 (95=F, 66=M), HIV = 204 (99=F, 105=M), Drug/toxin = 86 (53=F, 33=M)	1998-2022	Study on long term oxygen therapy; Authors - all PAH+CTEPH = 4533 (2926); CTEPH = 1364 (775=F, 598=M); Others note 52% (396) CHD has Eisenmenger.	[112]
FOCUS-PAH (6 Italian Hospitals, 1 UK)	410	59.4	n/a	66.0%	56.0%	IPAH = 155 (87=F); HPAH = 16 (11=F); CTD = 129 (114=F); CHD = 30 (22=F); PoPH = 51 (21=F); HIV = 17 (9=F); Drugs/toxins = 9 (6=F); Schistosomiasis = 1 (F)	April 2001 - Nov 2022	Long term follow up of 5 years; 2 out of 3 enrolled were female and associated with decreased longer term mortality compared with males	[4]
NPHU - National PH Unit (Ireland)	163	56	56	77.0%	79.0%	IPAH = 33 (26=F); HPAH = 2 (2=F); CTD = 80 (72=F); CHD^^ = 22 (13=F); PoPH =11 (2=F); HIV = 2 (2=F); PVOD = 5 (2=F); Drugs/toxin = 3 (3=F); HHT-PAH = 2 (1=F); CCB-PAH = 3 (3=F) long term calcium channel blocker responders	2010-2020	Group 4 - CTEPH = 67 (29=F, 43%) ^^Authors note CHD usually followed by cardiology	[113]
FINPAH (Finland)	268	57	n/a	73%	n/a	IPAH (n=106)*; HPAH (n=10)*; CTD (n=76)*; CHD (n=39); PoPH (n=7)*; HIV (< 5)*; PVOD (n=12)*; Drugs/toxin (n=7)	2008-2019	CTEPH (n=198)*; Other PH - Group 2, 3, 5, unclear (n= 45)	[114]
ASPIRE (UK)	385 (SSc-PAH only)	66.8	n/a	90% (SSc-PAH only)	n/a	PAH-SSc = 385 (347^=F)	2000-2020	SSc patients only -- Group 2 PH = 61 (90% F); Group 3 PH = 231 (75% F); unclassified PH = 39 (80% F)	[115]
ASPIRE (UK)	185	n/a	52	-	72%	Classical IPAH = 185 (133=F, 52=M); IPAH with lung phenotype = 139 (75=F, 64=M)	2001-2022	Classical IPAH only -- authors evaluate a larger group of patients with Group 3.1 or 3.2 PH = 375 (148=F, 227=M), and IPAH with lung phenotype	[116]
COMPERA (Official PH Registry of Germany, other countries participate)	128	n/a	45	-	77%	Classical IPAH = 128 (99=F); IPAH with lung phenotype = 268 (95=F, 173=M)	2007-2022	Classical IPAH only -- authors evaluate a larger group of patients with Group 3.1 or 3.2 PH = 910 (336=F, 574=M), and IPAH with lung phenotype
COMPERA (Official PH Registry of Germany, other countries participate)	2531	64.4	65.9 (I/H/D-PAH)	63.6%	59.8% (I/H/D-PAH)	I/H/D-PAH = 1698 (1016=F); CTD = 536 (434=F); CHD = 128 (84=F); HIV = 24 (11=F); PoPH =145 (64=F)	2010-2019	Large European registry; I/H/D-PAH 59.8% F; CTD 81% F; CHD 65.6% F and avg age 50.6 years; HIV = 45.8% F and avg age 45.8; PoPH = 44.1% F, avg age 57)	[117]
COMPERA (Official PH Registry of Germany, other countries participate)	1120	n/a	72	n/a	61.6%	All IPAH	Jan 2009 - Dec 2021	IPAH only -- authors evaluate IPAH and comorbidities; age ≥ 18 only; treatment naïve initially. IPAH pts with no comorbidities were younger, 67% F compared with 3-4 comorbidities group, which was older (74 avg age, 54.2% F)	[118]
Gi-PH-Reg / GIESSEN (Germany)	685	51	n/a	65%	n/a	IPAH = 294, 42.9% of PAH pts, CTD = 145, 21.2%; CHD = 91 13.3%; PoPH = 7.4%; PVOD 4.1%; HIV = 3.9%, other causes 7.2%	1993-2011	Registry with all groups of PH; CTEPH = 459 (258=F), 56% female; Most CHD had Eisenmenger (n=21)	[119]
Swiss Pulmonary Hypertension Registry	136	56	n/a	65%	n/a	IPAH = 68, HPAH = 0, drugs/toxins = 47; CTD = 12; CHD = 5; PoPH = 2; HIV = 2	2001-2005	Age < 18 excluded; CTEPH = 383 (181=F)	[120]
	190	58	n/a	57%	n/a	IPAH = 94, HPAH = 0, drugs/toxins = 40; CTD = 26; CHD = 15; PoPH = 4; HIV = 9	2006-2010
	165	58	n/a	61%	n/a	IPAH = 90, HPAH = 7, drugs/toxins = 9; CTD = 32; CHD = 12; PoPH = 8; HIV = 6	2011-2015
	69	65	n/a	58%	n/a	IPAH = 28, HPAH = 8, drugs/toxins = 2; CTD = 27; CHD = 1; PoPH = 3; HIV = 0	2016-2019
SPAHR (Swedish)	457	67	69 (I/HPAH)	64%	55% (I/HPAH)	I/HPAH = 227 (55% F); CTD = 140 (78% F); CHD = 61 (69% F); Other = 29 (52% F)	2000-2014	CTEPH = 183 (50% F), avg age 70. Avg age for CHD = 43	[121]
HOPE (Greece)	231	51.8	53.5 (I/HPAH)	64.5%	55.8%	IPAH/HPAH = 86 (48=F); CTD = 71 (56=F); CHD = 60 (37=F)	2015-April 2018	Registry launched in 2015, and also tracking CTEPH	[122]
Latvia	23	67.8	n/a	73.9%	n/a	IPAH = 15, CTD = 2; CHD = 4; PoPH = 1; HIV = 1	2019	Incidence of PAH in Latvia in 2019, age ≥ 18, diagnosis during 2019	[123]
Russian National Registry (Russia)	487	45.2	41	81.3%	84.4%	IPAH = 199 (168=F); CTD = 94 (84=F); CHD = 178 (133=F); Other PAH = 16 (11=F)	2012 - 2019	≥ 18 years old included; CTEPH = 206 (126=F), 61% F, avg age 52	[124]
BNP-PL Registry (Poland)	970	46.8	54.6 (I/HPAH)	69.8%	71.6% (I/HPAH)	I/HPAH = 444 (318=F); CTD = 132 (115=F); CHD = 356 (223=F); PoPH = 25 (15=F); Drugs/toxins = 7 (3=F); HIV = 6 (3=F)	n/a - 2018	Registry also includes CTEPH and pediatric CHD cases, not included here.	[125]
Registry	Asia
	Total number patients	Median age (years)		Female %		Subtypes PAH (if known)	Dates Represented	Other	Ref
		PAH	IPAH	PAH	IPAH
China	2031	35	35	76.2%	77.4%	IPAH = 788 (610=F); CTD = 266 (258=F); CHD = 917 (642=F); Others = 60 (37=F) -- HPAH = 38, Drug/toxin = 3; HIV = 6; PVOD/PCH = 6]	Aug 2009 - Aug 2019	Excluded CTEPH, age less ≤ 18	[126]
JAPHR (Japan)	316	47.9	n/a	76.3%	n/a	I/HPAH = 158; CTD = 78; CHD = 41; PoPH = 29; Other = 10	2008-2015	Authors examine differences in characteristics and treatment between two time periods; n = 39 PAH pts by RHC excluded due to no treatment	[127]
	315	52.7	n/a	79.4%	n/a	I/HPAH = 161; CTD = 102; CHD = 22; PoPH = 22; Other = 8	2016-2020
JAPHR (Japan)	129	45	n/a	73.6%	71.1%	IPAH = 45 (32=F); CTD = 41 (36=F); CHD = 31 (19=F); PoPH = 11 (7=F); Drugs/toxins = 1 (1=F)	April 1999 - Oct 2014	Sex differences in PAH patients in Japan -- female pts with better survival compared with male pts	[128]
KORPAH (Korea)	625	47.6	45.1	80.5%	73.1%	IPAH = 145 (106=F); APAH = 480 (397=F) with subgroups CTD = 311 (274=F); CHD = 159 (116=F); other = 10 (7=F)	Sept 2008 - Dec 2011	No significant differences between RHC for IPAH and APAH; mean 1.7 year follow up with n=35 mortality, and CTD-PAH had the highest mortality	[129]
PROKERALA (Kerala, India)	317	47.6	n/a	62.6%	n/a	IPAH represented 75% of group 1 PAH followed by: 22.4% CHD, CTD = 2.4%	n/a	ECHO and RHC used for diagnosis; Largest PH registry on the Asian subcontinent. ≥ 18 yo; Registry with all groups of PH. CTEPH = 73 (45=F)	[130]
COHARD-PH (Indonesia)	411 (CHD-PAH only)	36.4 (CHD-PAH)	n/a	81.8%	n/a	CHD-PAH = 411 (336=F, 75=M); CHD with no PH/PAH = 203 (161=F, 42=M)	2012-2019	Adult CHD registry only; Of the 614 RHC completed, 411 had PAH. 89% had ASD. No PAH = 203 (161=F, 42=M), avg age 32.2 yo, 81.8% had ASD. Complex CHD were excluded from this registry	[131]
Registry	Middle East
	Total number patients	Median age (years)		Female %		Subtypes PAH (if known)	Dates Represented	Other	Ref
		PAH	IPAH	PAH	IPAH
Iran	353 (including CTEPH)	48.6 (including CTEPH)	48.2	63.7% (including CTEPH)	66.0%	IPAH = 182 (121=F, 61=M), CHD (Eisenmenger) = 43 (30=F), CTD = 22 (20=F)	2009 – 2019	Total registry 353 (225=F, 125=M); CTEPH = 95 (50=F). There are n=11 not described	[132]
SAUDIPH (Saudi Arabia)	128	30.5	32 (I/HPAH)	74.2%	76.8% (I/HPAH)	I/HPAH = 56 (43=F); CTD = 14 (12=F); CHD = 53 (34=F); PoPH = 1; PVOD/PCH = 4	2004-2018	Registry with all groups of PH; CTEPH = 68 (46=F); Most CHD had Eisenmenger (n=21)	[133]
UAEPH (United Arab Emirates)	83	41	39	78.3%	79.0%	IPAH = 25 (19=F); CTD = 27 (25=F); CHD = 26 (18=F)	Jan 2015 - Dec 2021	Registry with all groups of PH; CTEPH = 34 (23=F)	[134]
THALES (Turkey)	1034	8.7	n/a	58.0%	n/a	All CHD-PAH; (49% Eisenmenger; 42.7% systemic to pulmonary shunts)	May 2009 - September 2011	Registry of PAH-CHD - pediatric and adult; 35% ≥ 18 years old	[135]
Registry	Other Regions
	Total number patients	Median age (years)		Female %		Subtypes PAH (if known)	Dates Represented	Other	Ref
		PAH	IPAH	PAH	IPAH
PHSANZ (New Zealand and Australia)	2044	55.1	54.5 (I/H/D PAH)	73% ^	72% ^	I/H/D PAH = 861 (2.8:1 F:M); CTD-PAH = 741 (5.2:1 F:M); CHD-PAH = 227 (2.5:1 F:M)	Jan 2004 - Dec 2011	3900 patients, 21 referral centers. CTD-PAH were 75% SSc-PAH. HPAH = 44; DPAH = 27; HIV = 12; PoPH = 61; Multifactorial = 124	[136]
RESPHIRAR (Southern Brazil)	370	41.8	45.1	78.5%	80.8%	IPAH = 125 (101=F); CTD = 88 (78=F); CHD = 116 (83=F)	2007-2017	Authors note that schistosomiasis-PAH is the 3rd leading cause of PAH in Brazil, but in the southern region (where registry is based), schistosomiasis is not endemic	[137]
RECOPILAR (Argentina)	399	47	n/a	78.7%	n/a	n/a	2014-2016	Group 1 PAH = 399 (78.7% F); Group 2 = 100 (40% F); Group 3 = 52 (51.9% F); Group 4 = 61 (68.9% F); Group 5 = 15 (73.3% F)	[138]
GSHPHR - Groote Schuur Hospital Pulmonary Hypertension Registry - (South Africa)	PAH = 26; Total PH = 58	n/a	n/a	79% (PAH + PH)	n/a	IPAH = 8; CTD = 6; CHD = 2; PoPH = 1; HIV = 9; Drug/toxin = 1 [F:M ratio unknown]	Oct 2015 - Nov 2017	88% of PAH pts completed RHC; Total PH patients = 58 (46=F, 79%), avg age 44 for All. Group 2 = 5, Group 3 = 5; Group 4 = 22, none in Group 5	[139]
PAPUCO (Pan-African)*** [Cameroon, Mozambique, Nigeria, & South Africa]	34	36	n/a	65%	n/a	Group 1 PAH = 34 (22=F); Echocardiographic diagnosis only (no RHC). Median RVSP 71 mmHg, median TAPSE 15mm	2011-2013	*** Echocardiographic dx, no RHC. PH Registry of 220 consecutive PH pts (209 adult, 11 children); Group 2 = 144 (85=F, 59%, avg age 53); Group 3 = 23 (10=F, 44%, avg age 43)	[140]

Key

NIH = National Institute of Health

PPH = Primary Pulmonary Hypertension

PAH = Pulmonary Arterial Hypertension

PH = Pulmonary Hypertension

HPAH = Heritable PAH

IPAH = Idiopathic PAH

APAH = Associated PAH

CTD = Connective Tissue Disease

CHD = Congenital Heart Disease

PoPH = Portopulmonary hypertension

HIV = Human Immunodeficiency Virus

PVOD = Pulmonary Veno-occlusive disease

D-PAH = Drug/Toxin associated PAH

PCH = Pulmonary Capillary Hemangiomatosis

CTEPH = Chronic thromboembolic pulmonary hypertension

SSc = Systemic Sclerosis

BNP = Brain Natriuretic Peptide

NT-proBNP = N-Terminal Pro-Brain Natriuretic Peptide

WHO-FC = World Health Organization - Functional Class

6MWD = Six-minute walk distance

TTE = Transthoracic echocardiogram

RHC = Right heart catheterization

ASD = Atrial septal defect

HHT = hereditary hemorrhagic telangiectasia associated PAH

CCB = Calcium channel blocker

^{^}= our calculation from author data

^{^^}= authors note that CHD patients usually follow with cardiology

^**= No sex data

M = Male

F = Female

pts = patients

^***Registry without RHC as a requirement - authors note resource limitation

The Right Ventricle and Pulmonary Artery Coupling

In epidemiologic cohorts without cardiovascular disease, women have higher RV ejection fractions, lower RV volumes and mass as compared to men and after adjustment for anthropometrics and left ventricular function [79,80]. Sex steroid levels and sex hormone genotypes (variation in the CYP1B1 gene) have been linked to RV phenotype in the Multi-Ethnic Study of Atherosclerosis epidemiologic cohort. Specifically, higher levels of circulating E2 were associated with higher RV ejection fraction and lower RV volumes in postmenopausal women using hormone therapy; polymorphisms in the CYP1B1 gene linked to RV function were also tightly linked to the polymorphism identified to increase PAH penetrance in HPAH [81,82].

As in healthy individuals, there are sex differences in RV remodeling and adaptation in PAH. Women with variable types of pulmonary vascular disease are known to have better RV function than men; this has been shown in PAH, in chronic left heart and lung diseases, and possibly CTEPH, and reviewed elsewhere [83-85*]. In a retrospective cohort study of 101 idiopathic PAH patients, male sex was an independent predictor of worse survival due to deterioration of RV function [86]. In a follow-up study by the same group, sex hormone levels had sex-specific relationships with RV function [65]. 17β-estradiol has been demonstrated to restore pulmonary artery (PA) compliance, reduce pulsatile loading, and improve ventricular-vascular coupling in PH models [87]. In clinical trial participants with PAH, older females had significantly higher PA compliance than younger females but overall lower PA compliance as compared to males [76]. These findings differ from Tello et al, which showed that better PA – RV coupling in females with PAH, irrespective of age [88]. In a recently published PA banding model of RV failure, investigators found that male mice developed more severe RV dysfunction and failure, marked by maladaptive RV remodeling with increase in RV collagen deposition, degradation of the extracellular matrix, and recruitment of macrophages. Female mice lacked chronic inflammation and had less severe RV dysfunction [89**]. Additional mechanistic research is needed to identify potential therapeutic targets for sex-based differences in RV inflammation and remodeling.

A sex-stratified genome-wide association study from the UK Biobank identified a potential candidate gene – BMPR1A – for female specific genetic determination of RV function in PAH. Variation in BMPR1A in females with HPAH and idiopathic PAH was also associated with higher CI [90**]. Embryonically, BMPR1A contributes to RV growth, and selective defects in BMPR1A are associated with endocardial cushion defects, conduction delays, and other cardiac defects. Interestingly, BMPR1A is a receptor for anti-Müllerian hormone (AMH), and declining AMH levels have been associated with cardiovascular health in women [91].

Response to therapy

There are sex differences in response to therapy in PAH. In part, this may be due to physiological and hormonal differences between males and females that impact the cellular milieu and intersect with specific PAH therapies. Plasma concentrations of ET-1 are higher in males compared with females, and ET-1 concentrations increase with age [92]; endothelial receptors A and B (ET_A and ET_B) also vary by sex and change with age [93]. The loss of E2 with the menopausal transition not only impacts ET-1 but also enhances NO and prostacyclin (PGI₂) [94], which may explain attenuation of sex-based phenotypes with age in some PAH studies. Observational data and post hoc analyses of PAH clinical trials indicate differential responses to PAH therapeutic classes by sex or gender. In the Spanish PAH registry (REHAP) and the PHIRST randomized clinical trial of tadalafil, men were more likely to respond to phosphodiesterase-5 inhibitors (PDE5i) [95*,96]. In a pooled analysis of 1130 PAH participants from clinical trials, women were more likely to improve on endothelin receptor antagonists (ERAs) than men [97].

Behaviors, Social Determinants of Health, and Intersectionality

We have discussed chromosomal, genetic, and hormonal factors that may explain the observed “sex paradox” in PAH. In addition to these biological influences, there are societal, cultural, and political constructs that may also be gendered and influence prevalence and outcomes in PAH. In a recent study from the Pulmonary Hypertension Association Registry (PHAR), male sex, in addition to poverty, being unpartnered, being underinsured, and less educated were all risk factors for PAH medication non-adherence [98*], which was associated with more emergency room visits, hospitalizations, and worse health-related quality of life. Collider bias, additional variables that may directly influence sex and outcomes creating the counterintuitive “sex paradox,” may also play a role. In another recent study from the PHAR, despite worse hemodynamics, 6MWD, functional status and more PAH-therapy in women, women had a 48% lower risk of death compared with men. This survival difference was not mediated by high-risk behaviors, suggesting additional unmeasured factors may influence sex-based outcomes in PAH [2*]. Men with PAH are more likely to report tobacco and methamphetamine use compared with women, and these habits are independently associated with mortality in PAH [2*,99*,100]. Interestingly, more women develop methamphetamine associated-PAH while more men develop methamphetamine-induced cardiomyopathy [101]. Finally, PAH may impact health-related quality of life differently in men versus women; women with PAH report high rates of sexual dysfunction that may vary by PAH treatments [102]. As PDE5i are also approved for erectile dysfunction, males with PAH treated with PDE5i may have off-target effects, although this remains speculative. Finally, restrictions in access to reproductive care and expert PH care centers may have additional implications for individuals who are pregnant or seeking reproductive counseling [103*].

Conclusion

PAH exhibits marked sexual dimorphism with a global female predominance yet improved survival. Ongoing research continues to elucidate the complex interactions among biologic sex, genetics, steroidogenic sex hormones, their metabolites and receptors, inflammation and immune regulation, epigenetics and gender/intersectionality. While murine models of PH fail to fully recapitulate the intricacies of human PAH, they offer emerging mechanistic insights into the influence of sex chromosomes and hormones. As global PAH registry efforts expand and clinical trials that target (or affect) sex hormones are conducted, it is imperative to systemically capture data on biological sex, gender identity, hormonal exposures, social determinants of health and lifestyle factors. Such granularity will be essential for advancing our understanding of the sex paradox or puzzle of pulmonary vascular disease.

KEY POINTS:

• The “sex paradox” means that female sex is associated with the risk of pulmonary vascular disease, but females have improved right ventricular function and survival compared with males.

• Pathobiological pathways and mechanisms in PAH all intersect with sex hormone signaling and metabolism; recent work also suggests sex chromosomes may play a role in modulating pulmonary vascular disease.

• Pro-inflammatory cytokines, immune function, and immune dysregulation interface with sex hormone metabolism and signaling and play an important role in PAH susceptibility and pathogenesis.

• Recently completed clinical trials on sex hormone modulation offer insight into the complexity of PAH.

• Sex and gender differences in behaviors and social determinants of health are also important factors in PAH and outcomes.