Transient Left Ventricular Dysfunction in Loeys-Dietz Syndrome With Loss-of-Function TGFBR1 Variants

Abstract

Background

Loeys-Dietz syndrome (LDS) is an autosomal dominant connective tissue disorder characterized by vascular and valvular diseases. TGFBR1 is a major causative gene, mediating disease progression through dysregulated transforming growth factor-β signaling; however, its role in left ventricular systolic function remains unclear.

Case Summary

We report 2 LDS cases harboring TGFBR1 variants (p.Asp400Gly and p.Leu196Pro). Severe left ventricular systolic dysfunction was observed in these patients but improved with initiation and optimization of medical therapy. Histopathologic analysis of right ventricular biopsy specimens revealed limited interstitial and perivascular fibrosis. Functional analysis demonstrated reduced transforming growth factor-β signaling activity associated with both variants and decreased protein expression of TGFBR1-Asp400Gly.

Discussion

These findings suggest a potential loss-of-function effect of TGFBR1 variants in LDS, and the clinical course including transient cardiac dysfunction provides important insights into future medical management for LDS.

Take-Home Message

Recognition of transient cardiac dysfunction is important for tailored heart failure management in LDS patients with TGFBR1 variants.

Visual Summary

Loeys-Dietz syndrome (LDS) is an autosomal dominant connective tissue disorder characterized by features such as aortic aneurysms, dissections, and valvular diseases that resemble those observed in Marfan syndrome. Patients with LDS are more prone to aortic dissection or rupture at a younger age and with smaller aneurysms compared with patients with Marfan syndrome, highlighting the importance of early surveillance.¹ LDS and the causative genes are classified into type I (TGFBR1), type II (TGFBR2), type III (SMAD3), type IV (TGFB2), and type V (TGFB3). In patients with type I LDS caused by the TGFBR1 variant, aortic aneurysms, abnormal uvula, arterial tortuosity, and arachnodactyly are the most frequently observed clinical characteristics.²

In addition to vascular manifestations, impaired left ventricular systolic function that cannot be fully accounted for by concomitant vascular diseases has been reported in rare cases of LDS with a TGFBR1 variant. A 44-year-old man with LDS who underwent emergent ascending aortic replacement and resuspension of the aortic valve due to acute aortic dissection developed heart failure with impaired systolic function (left ventricular ejection fraction [LVEF]: 25%). LVEF was improved to 40% with medical therapy, but the patient finally underwent heart transplantation due to decreased LVEF.³ A 50-year-old man with LDS developed heart failure with decreased LVEF (30%) and left ventricular dilatation (73 mm). After medical therapy, LVEF was improved to 48% and left ventricular diameter was decreased to 57 mm.⁴

Although LDS has been recognized as a disorder characterized by vascular abnormalities, clinical findings showing the association between LDS and left ventricular systolic dysfunction are limited. Here we report 2 patients with LDS harboring TGFBR1 variants (Asp400Gly and Leu196Pro) who developed severe but transient left ventricular systolic dysfunction after optimization of medical therapy.

Case Series

Patient 1

Patient 1 was a Japanese man with no apparent family history of cardiovascular disease, including aortic aneurysm, aortic dissection, arterial tortuosity, or valvular heart disease (Figure 1A). At 19 years of age, he was hospitalized for pneumothorax. Because of multiple comorbidities, including scoliosis and atlantoaxial subluxation, genetic testing was performed, and a pathogenic variant in TGFBR1 (NM_004612.4 [NP_004603.1]: c.1199A>G, p.Asp400Gly) was identified (Figure 1B). Based on the clinical symptoms and genetic analysis,¹ he was diagnosed with LDS. At 21 years of age, he was hospitalized for subarachnoid hemorrhage and underwent regular cardiovascular follow-up after discharge. At 26 years of age, routine echocardiography revealed a reduced LVEF of approximately 40% and severe mitral regurgitation (MR) (Figure 1C). He subsequently underwent minimally invasive mitral valve repair, resulting in improvement of MR to a trivial degree. At the same age, he was hospitalized for an acute subdural hematoma. Seven months after MR surgery, transthoracic echocardiography detected progressive left ventricular dilatation and systolic dysfunction (left ventricular end-diastolic diameter/left ventricular end-systolic diameter: 63/54 mm; LVEF: 19%). His heart failure symptoms gradually worsened. At 27 years of age, poor compliance with optimal medical therapy resulted in further left ventricular dilatation (left ventricular end-diastolic diameter/left ventricular end-systolic diameter: 74/70 mm; LVEF: 17%), requiring hospitalization. During hospitalization, guideline-directed medical therapy was optimized, including sacubitril/valsartan, bisoprolol, spironolactone, dapagliflozin, furosemide, and ivabradine, and follow-up transthoracic echocardiography demonstrated an improvement in LVEF compared with the previous assessment. He was discharged after stabilization of heart failure, but exertional dyspnea persisted and was consistent with NYHA functional class II symptoms. At 28 years of age, during outpatient follow up, LVEF recovered to 56% (Figure 1D).

Figure 1.

Clinical Course of 2 Patients With Loeys-Dietz Syndrome With TGFBR1 Variants Exhibiting Left Ventricular Dysfunction

(A) Pedigrees of the probands. Individuals affected with Loeys-Dietz syndrome indicated by filled circles (females) or filled squares (males). In patient 1, a TGFBR1 variant (TGFBR1-Asp400Gly) was identified in the proband indicated by an arrow (II-2). In patient 2, a TGFBR1 variant (TGFBR1-Leu196Pro) was identified in the proband indicated by an arrow (I-2) and her daughter (II-2). (B) Results of Sanger sequencing analysis of TGFBR1 after polymerase chain reaction amplification using genomic DNA extracted from the peripheral blood of the patient 1 proband. (C) Mitral regurgitation detected by transthoracic echocardiography in patient 1 at 27 years of age (apical 4-chamber view with Doppler imaging). (D) Longitudinal changes in LVEF and clinical course in patient 1. The x-axis indicates the patient's age (in years). Between the ages of 26 and 27, medication compliance in patient 1 was inadequate. (E) Longitudinal changes in LVEF and clinical course in patient 2. The x-axis indicates the patient's age (in years). ARB = angiotensin receptor blocker; ARNI = angiotensin receptor neprilysin inhibitor; hANP = human atrial natriuretic peptide; LVEF = left ventricular ejection fraction; MICS-MVP = minimally invasive cardiac surgery–mitral valve plasty; MRA = mineralocorticoid receptor antagonist; SGLT2i = sodium-glucose cotransporter 2 inhibitor; TEVAR = thoracic endovascular aortic repair.

Patient 2

Patient 2 was a Japanese woman treated under a clinical diagnosis of Takayasu arteritis. At 34 years of age, she underwent graft replacement for an abdominal aortic aneurysm and subsequently required multiple additional aortic surgeries over the following decade. Her daughter was previously diagnosed with LDS carrying a TGFBR1 variant (NM_004612.4 [NP_004603.1]: c.587T>C, p.Leu196Pro) (Figure 1A), and genetic analysis clarified that she carried the same TGFBR1 variant, confirming LDS at 48 years of age. At 59 years of age, she was admitted for exertional dyspnea due to heart failure. Transthoracic echocardiography revealed left ventricular dilatation and reduced systolic function (left ventricular end-diastolic diameter: 51 mm; LVEF: 38%). Her symptoms improved with optimized medical therapy, and she was discharged. At 60 years of age, she was readmitted with acute decompensated heart failure and new-onset atrial tachycardia. Her LVEF had declined to 22%, and she required inotropic support and rhythm control. Her hospital course was prolonged because of difficulty controlling heart failure in the setting of gastrointestinal bleeding. Volume reduction was achieved using human atrial natriuretic peptide, after which the dose of an angiotensin receptor blocker was increased. Subsequently, angiotensin receptor neprilysin inhibitor therapy was initiated. With continued management, her cardiac function gradually recovered, and follow-up evaluation at age 61 showed an improvement in LVEF to 67% (Figure 1E).

Histopathologic analysis

Both patients underwent right ventricular endomyocardial biopsy evaluation to examine the pathologic basis underlying the severely reduced contractile function. Representative histopathologic images of right ventricular myocardial biopsies from patient 1 (TGFBR1-Asp400Gly) (Figure 2A) and patient 2 (TGFBR1-Leu196Pro) (Figure 2B) are shown. In patient 1, most of the myocardial tissue was viable, with minimal evidence of interstitial or perivascular fibrosis. There was no apparent increase in elastic fibers. In patient 2, nuclear enlargement and deformation were observed in cardiomyocytes. In addition, a subset of cardiomyocytes exhibited basophilic degeneration and vacuolar changes in the cytoplasm. Mild interstitial and perivascular fibrosis were present in limited areas (Table 1). These data suggest that fibrotic changes were not apparent in myocardial histopathologic specimens in either patient 1 or patient 2, although both patients exhibited reduced ventricular contraction.

Figure 2.

Histopathologic Analysis Revealed Minimal Fibrosis in the Right Ventricular Myocardium of 2 Patients With Loeys-Dietz Syndrome With TGFBR1 Variants

(A) Patient 1: hematoxylin and eosin staining, scale bar: 100 μm. Masson's trichrome staining was not available because of limited clinical material. (B) Patient 2: hematoxylin and eosin staining (top) and Masson's trichrome staining (bottom), scale bar: 200 μm.

Table 1.

Histopathologic and Functional Findings

Finding	Patient 1	Patient 2
Right ventricular endomyocardial biopsy (fibrosis)	Viable myocardium; minimal interstitial and perivascular fibrosis; no apparent increase in elastic fibers	Nuclear enlargement; basophilic degeneration; mild and limited interstitial/perivascular fibrosis
In vitro transforming growth factor-β signaling (HEK293T)	Reduced SMAD-responsive luciferase activity; reduced TGFBR1 protein expression	Reduced SMAD-responsive luciferase activity

Functional analysis

To investigate the functional impact of TGFBR1 variants on TGF-β signaling, we examined receptor activity using HEK293T cells (Supplemental Methods). TGF-β stimulation dose-dependently increased luciferase activity in HEK293T cells expressing TGFBR1-WT, and these activities were significantly reduced in cells expressing TGFBR1-Asp400Gly or Leu196Pro, suggesting reduced responsiveness of TGFBR1 to TGF-β stimulation (Figure 3A). The reduced TGF-β activity in the Asp400Gly variant was consistent with the previous findings demonstrating reduced reporter activities in genetic variants in TGFBR1 detected in LDS patients.⁵ Next, we evaluated protein expression levels using HEK293T cells transfected with TGFBR1-WT, -Asp400Gly, or Leu196Pro and found that protein expression levels of TGFBR1-Asp400Gly were significantly reduced compared with TGFBR1-WT (Figure 3B). The functional characteristics of the 2 TGFBR1 variants analyzed in this study (Table 1), together with previously reported TGFBR1 variants identified in LDS patients, are summarized in Supplemental Table 1. All listed variants showed reduced SMAD-responsive luciferase activity and were classified as loss-of-function. Supplemental Figure 1 summarizes the domain architecture of TGFBR1 and the locations of the 2 TGFBR1 variants analyzed in this study and previously reported variants. Almost all variants including TGFBR1-Asp400Gly were located within the kinase domain, and TGFBR1-Leu196Pro and Thr200Ile variants were located within the glycine-serine–rich domain, which are critical for receptor activation and downstream SMAD signaling.⁶

Figure 3.

TGFBR1-Asp400Gly and Leu196Pro Reduced TGF-β Signaling Activities

(A) HEK293T cells were co-transfected with SBE4-Luc and expression vectors encoding WT or variant TGFBR1. Twenty-four hours after vehicle or TGF-β1(10, 50, 100 ng/mL) stimulation, cells were collected and luciferase activity was measured. Data are presented as relative luciferase activity normalized to the activity of vehicle control, which was set to 1. Data are shown as mean ± SD. Statistical significance was assessed by 2-way analysis of variance followed by Dunnett's multiple comparisons test. ∗P < 0.0001. (B) HEK293T cells were transfected with expression vectors encoding WT or variant TGFBR1, and protein expression was evaluated by Western blotting. Representative Western blots (left) and quantitative analysis shown as bar graphs (right) are presented. TGFBR1 protein levels were quantified by densitometric analysis of Western blot bands and normalized to GAPDH. Relative TGFBR1 protein expressions are expressed relative to WT, which was set to 1. Data are presented as mean ± SD (n = 4). Statistical analysis was performed using 1-way analysis of variance followed by Dunnett's multiple comparisons test with WT as the control. ∗P < 0.05 vs WT. GAPDH = glyceraldehyde-3-phosphate dehydrogenase NC = non-transfected control; NS = not significant; TGF-β; transforming growth factor-β; WT = wild-type.

Discussion

In this study, we reported 2 patients with LDS with TGFBR1 variants and concurrently exhibited left ventricular dysfunction. Although LVEF values evaluated by echocardiography in both patients were substantially decreased (19% in patient 1, 22% in patient 2), LVEFs were gradually recovered after optimization of medical therapy. Characteristic clinical courses in the current cases combined with the previously reported LDS cases^3,4 may imply the underlying nature of left ventricular systolic dysfunction in LDS. In patient 1, left ventricular dysfunction became apparent concomitant with severe MR at 26 years of age, and the LVEF gradually worsened although MR was improved to a trivial degree by surgical operation. Decrease in LVEF after mitral valve repair is potentially caused by removal of the excess preload and increased afterload in ventricle.⁷ During this period, poor compliance with the indicated medical therapy combined with hemodynamic alterations resulted in further decrease in LVEF in patient 1. Patient 2 developed ventricular dysfunction after a certain interval after multiple endovascular interventions, including stent-graft implantation such as endovascular aneurysm repair and thoracic endovascular aortic repair. In endovascular therapy, stent-graft implantation reduces aortic compliance and elasticity, leading to increased left ventricular afterload and similar hemodynamic effects.⁸ In the present 2 cases, it is possible that pre-existing vulnerability due to molecular abnormalities, combined with surgical invasiveness and hemodynamic changes, contributed to the manifestation of left ventricular dysfunction.

The pathogenic effects of TGF-β signaling in the heart are believed to be mediated primarily through cardiac fibroblasts. Consistent with this notion, neutralization of TGF-β in pressure overload models has been shown to attenuate myocardial fibrosis and improve diastolic function,⁹ suggesting that TGF-β signaling plays a central role in cardiac interstitial fibrosis. However, fibroblast-specific inhibition of TGF-β signaling has also been reported to reduce fibrosis without necessarily improving chamber dilatation or cardiac dysfunction.¹⁰ These findings indicate that suppression of fibrosis does not invariably translate into functional improvement. Under conditions of pressure overload, a certain degree of fibrosis may contribute to the maintenance of myocardial structural integrity and stiffness. Thus, insufficient fibrosis could potentially impair adaptation to increased afterload. In both cases examined in the present study, cardiac interstitial fibrosis was relatively mild, raising the possibility that inadequate fibrotic remodeling associated with abnormal TGF-β signaling may alter the biomechanical properties of the myocardium and reduce its ability to adapt to increased afterload. Under these conditions, the additional surgical or interventional treatments may trigger the development of left ventricular dysfunction.

Our in vitro analyses demonstrated the decreased TGF-β activity in cells expressing the TGFBR1 variant. TGFBR1 consists of multiple functional domains, including the glycine-serine–rich domain and the kinase domain. The glycine-serine–rich domain is phosphorylated by TGFBR2 and plays a role in receptor activation, whereas the kinase domain mediates phosphorylation of SMAD2/3.⁵ Examination of previously reported functional analyses indicates that many pathogenic TGFBR1 variants are clustered within the kinase domain (Supplemental Table 1, Supplemental Figure 1), highlighting the importance of this region in TGF-β signaling activity. In the present study, TGFBR1-Asp400Gly was located near the catalytic loop within the kinase domain, whereas TGFBR1-Leu196Pro was located near the glycine-serine–rich domain. Previous studies have demonstrated that the catalytic loop and surrounding residues form essential structural units required for kinase activity, and variants in this region may affect the catalytic function of TGFBR1.⁵ The TGFBR1-Asp400Gly variant showed reduced protein expression levels, suggesting that variants located in structurally and functionally critical regions may impair receptor stability and affect the formation of the TGFBR1–TGFBR2 complex.

In conclusion, we report 2 patients with LDS with left ventricular dysfunction associated with TGFBR1 variants. Our in vitro analyses demonstrated reduced TGF-β signaling, and these alterations may lead to changes in the mechanical properties of the myocardium and interstitial tissue. To clarify the impact of TGFBR1 variants on cardiac function, further studies are warranted that focus on the mechanical properties of cardiomyocytes and interstitial tissue.

Funding Support and Author Disclosures

This research was supported by JSPS KAKENHI (24K11239, 24K02446, 24K22121), Fusion-Oriented Research for Disruptive Science and Technology (J250701505), Health and Labor Sciences Research grant (23FC1050), Japan Heart Foundation, the Center for Medical Innovation and Translational Research, and the Center for Medical Research and Education at the Graduate School of Medicine, The University of Osaka. The authors have reported that they have no relationships relevant to the contents of this paper to disclose.