The first case report of de novo Crohn's disease after heart transplantation successfully treated with ustekinumab.

Inflammatory bowel disease (IBD) is a complex chronic inflammatory intestinal disease. The development of de novo IBD after solid organ transplantation with immunosuppressive agents has been rarely reported. We present the case of a 65-year-old man with repeated colitis after heart transplantation (HTx) who was diagnosed with Crohn's disease (CD). The patient underwent HTx due to non-ischemic dilated cardiomyopathy. Six months after HTx, he developed serious diarrhea and a transient fever, which persisted for about 6 months. Valganciclovir or any antibiotic agents were not effective for his symptoms and longitudinal ulcers in colonoscopy aggravated during the course, so that we made a diagnosis of CD. We started 5-aminosalicylic acid and found improvement in his symptoms and colonoscopic findings. However, 7 months after improvement, CD worsened. We started ustekinumab by which his condition successfully went into remission again. While oral immunosuppressive drugs are thought to suppress autoimmune diseases in general, IBD should be included in the differential diagnoses for recurring enterocolitis after HTx. Poorly controlled CD can lead to serious and potentially fatal complications, but in this case, ustekinumab has been used safely and effectively for the treatment of CD. Learning objective: Colitis is a common complication after heart transplantation (HTx). Although cytomegalovirus colitis or posttransplant lymphoproliferative disorder are observed commonly, de novo inflammatory bowel disease (IBD) should be considered when serious refractory colitis occurs. Not only 5-aminosalicylic acid but also ustekinumab, which is a monoclonal antibody to the p40 subunit of interleukin (IL)-12 and IL-23, may be a safe and effective treatment for de novo IBD after HTx.

Comparison of osteoanabolic agents (teriparatide and romosozumab) with bisphosphonates in prevention of subsequent vertebral fractures in patients treated for osteoporotic vertebral fracture for 12 months: An observational cohort study.

Introduction: Domino osteoporotic vertebral fracture (OVF) is as a subsequent fracture that develops within 3 months before the initial OVF heals. There is limited evidence regarding the efficacy of osteoanabolic agents on its treatment. This study evaluated the effects of bisphosphonates and anabolic agents teriparatide and romosozumab on subsequent domino OVF. Methods: This was post hoc analysis of a prospective, multicenter, observational study conducted across 8 hospitals, enrolling 144 patients with conservatively treated OVF, grouped into patients receiving bisphosphonate (BP, n = 55), teriparatide (TPTD, n = 62), and romosozumab (Romo, n = 27). The primary outcome was the incidence of subsequent OVF at 3 and 12 months, whereas the secondary outcomes included the incidence of pseudoarthrosis and progression of vertebral collapse (VC). Pseudoarthrosis was classified as stable or unstable based on vertebral instability. Results: The use of osteoanabolic agents did not reduce the incidence of subsequent OVF at 3 and 12 months. There were no significant differences in the background data or type of conservative treatment among the three groups. However, the TPTD and Romo groups had significantly lower rates of unstable pseudarthrosis (p = 0.03). Additionally, there were no significant differences in VC progression between groups, but it tended to be higher in the BP group than the TPTD and Romo group (p = 0.07). Conclusion: Osteoanabolic agents were beneficial in reducing unstable pseudoarthrosis, but were not more effective than bisphosphonates in the development of subsequent domino OVF. A more comprehensive approach to the treatment of osteoporosis is needed to prevent domino OVFs.

Right heart reserve function assessed with fluid loading predicts late right heart failure after left ventricular assist device implantation.

BACKGROUND: A left ventricular assist device (LVAD) is an effective therapeutic option for advanced heart failure. Late right heart failure (LRHF) is a complication after LVAD implantation associated with increasing morbidity and mortality; however, the assessment of right heart function, including right heart reserve function after LVAD implantation, has not been established. We focused on a fluid loading test with right heart catheterization (RHC) to evaluate right heart pre-load reserve function and investigate its impact on LRHF. METHODS: Patients aged > 18 who received continuous-flow LVAD between November 2007 and December 2022 at our institution and underwent RHC with saline loading (10ml/kg for 15 min) 1 month after LVAD implantation were included. RESULTS: Overall, 31 LRHF or deaths (RHF group) have occurred in 149 patients. Comparing the RHF and non-RHF groups, pulmonary artery pulsatility index (PAPi) at rest (1.8±0.89 vs. 2.5±1.4, p=0.02) and right ventricular stroke work index (RVSWi) change ratio with saline loading (0.96±0.32 vs. 1.1±0.20, p=0.03) was significantly different. The PAPi at rest and RVSWi change ratio with saline loading were identified as the postoperative risks for LRHF or death. The cohort was divided into three groups based on whether the PAPi at rest and RVSWi change ratio were low. The event-free curve significantly differed between the three groups (p<0.001). CONCLUSIONS: Hemodynamic assessment with saline loading can evaluate the right ventricular pre-load reserve function of patients with LVAD. The low RVSWi change with saline loading was a risk factor for LRHF following LVAD implantation.

Comprehensive Risk Assessment in Patients With Pulmonary Arterial Hypertension Referred for Lung Transplantation.

BACKGROUND: Whether comprehensive risk assessment predicts post-referral outcome in patients with pulmonary arterial hypertension (PAH) referred for lung transplantation (LT) in Japan is unknown.Methods and Results: We retrospectively analyzed 52 PAH patients referred for LT. Risk status at referral was assessed using 3- and 4-strata models from the 2022 European Society of Cardiology and European Respiratory Society guidelines. The 3-strata model intermediate-risk group was further divided into 2 groups based on the median proportion of low-risk variables (modified risk assessment [MRA]). The primary outcome was post-referral mortality. During follow-up, 9 patients died and 13 patients underwent LT. There was no survival difference among 3-strata model groups. The 4-strata model classified 33, 16, and 3 patients as low intermediate, high intermediate, and high risk, respectively. The 4-strata model identified high-risk patients with a 1-year survival rate of 33%, but did not discriminate survival between the intermediate-risk groups. The MRA classified 15, 28, 8, and 1 patients as low, low intermediate, high intermediate, and high risk, respectively. High intermediate- or high-risk patients had worse survival (P<0.001), with 1- and 3-year survival rates of 64% and 34%, respectively. MRA high intermediate- or high-risk classification was associated with mortality (hazard ratio 12.780; 95% confidence interval 2.583-63.221; P=0.002). CONCLUSIONS: Patients classified as high intermediate or high risk by the MRA after treatment should be referred for LT.

A case of progressive right ventricular failure with ventricular arrhythmia and aortic insufficiency after implantable left ventricular assist device implantation.

Right ventricular failure (RVF) is a serious complication after left ventricular assist device (LVAD) implantation. In this report, a case of RVF that developed over two years after LVAD implantation is presented. The patient was a 12-year-old male with dilated phase of hypertrophic cardiomyopathy. He had no risk factors for early or late-onset RVF. However, his right ventricular function worsened after he developed ventricular arrhythmia (VA), and right ventricular dysfunction became exacerbated with an increasing frequency of VAs. He also developed moderate aortic insufficiency (AI), which became severe. Two years after implantation, he was admitted for treatment of recurrent ventricular tachycardia and became inotropic-dependent during hospitalization. Finally, he underwent successful heart transplantation 2 years and 9 months after LVAD implantation. This case suggests that vicious cycle of RV dysfunction, recurrent VAs and severe AI could lead to RVF in patients without known risk factors for RVF, even long after LVAD implantation. Learning objective: This report shows a progressive right ventricular failure (RVF) two years after left ventricular assist device (LVAD) implantation. Although the patient had no known risk factor, vicious circle of RV dysfunction, ventricular arrhythmias (VAs) and aortic insufficiency (AI) lead to RVF. Patients with LVAD as destination therapy will increase and require long-term LVAD management. We should recognize that these patients could develop RVF even years after LVAD implantation in association with VAs and AI.

Pre-Transplant Seroprevalence, Associated Factors, and Post-Transplant Incidence of Toxoplasma gondii Infection Among Heart Transplant Recipients in Japan.

BACKGROUND: Among solid organ transplant (SOT) recipients, heart transplant (HT) recipients are at a higher risk of Toxoplasma gondii infection. As Toxoplasma seroprevalence varies by geographic location, updated local epidemiology is essential to guide preventive and therapeutic strategies. However, the Toxoplasma seroprevalence and incidence of post-transplant toxoplasmosis among SOT recipients in Japan are unknown. METHODS: We performed a single-center retrospective observational study at an HT center in Tokyo, Japan. All HT recipients aged ≥18 years between 2006 and April 2019 were included. We reviewed patient charts and conducted a questionnaire survey to investigate the risk factors for infection. RESULTS: Among 105 recipients included in the study, 11 (10.5%) were seropositive before transplant. Ninety-five recipients (90.5%), including all pre-transplant seropositive recipients, answered the questionnaire. The recipients who had lived in Okinawa (odds ratio [OR] 7.5 [95% CI 1.42-39.61]; P = .032) and who reported raw-meat eating habits (OR 4.64 [95% CI 1.04-23.3]; P = .021) were more likely to be seropositive. None of the patients developed symptoms of toxoplasmosis. The post-transplant incidence of other major adverse outcomes was not significantly different according to the pre-transplant serostatus. CONCLUSIONS: About 10% of HT recipients at an HT center in Tokyo were seropositive for Toxoplasma pre-transplant, and none developed symptomatic toxoplasmosis post-transplant on trimethoprim-sulfamethoxazole. The history of raw meat consumption was associated with seropositivity; therefore, avoiding it might be recommended for HT recipient candidates.

Myocardial DNA Damage Predicts Heart Failure Outcome in Various Underlying Diseases.

BACKGROUND: Reliable predictors of treatment efficacy in heart failure have been long awaited. DNA damage has been implicated as a cause of heart failure. OBJECTIVES: The purpose of this study was to investigate the association of DNA damage in myocardial tissue with treatment response and prognosis of heart failure. METHODS: The authors performed immunostaining of DNA damage markers poly(ADP-ribose) (PAR) and γ-H2A.X in endomyocardial biopsy specimens from 175 patients with heart failure with reduced ejection fraction (HFrEF) of various underlying etiologies. They calculated the percentage of nuclei positive for each DNA damage marker (%PAR and %γ-H2A.X). The primary outcome was left ventricular reverse remodeling (LVRR) at 1 year, and the secondary outcome was a composite of cardiovascular death, heart transplantation, and ventricular assist device implantation. RESULTS: Patients who did not achieve LVRR after the optimization of medical therapies presented with significantly higher %PAR and %γ-H2A.X. The ROC analysis demonstrated good performance of both %PAR and %γ-H2A.X for predicting LVRR (AUCs: 0.867 and 0.855, respectively). There was a negative correlation between the mean proportion of DNA damage marker-positive nuclei and the probability of LVRR across different underlying diseases. In addition, patients with higher %PAR or %γ-H2A.X had more long-term clinical events (PAR HR: 1.63 [95% CI: 1.31-2.01]; P < 0.001; γ-H2A.X HR: 1.48 [95% CI: 1.27-1.72]; P < 0.001). CONCLUSIONS: DNA damage determines the consequences of human heart failure. Assessment of DNA damage is useful to predict treatment efficacy and prognosis of heart failure patients with various underlying etiologies.

Multivessel Coronary Artery Dissection in a Patient with Co-Occurrence of Aortic Dissection and Dilated Cardiomyopathy in the Postpartum Period.

The co-occurrence of dilated cardiomyopathy (DCM) and aortic dissection has been rarely reported. Here, we present the case of a patient with co-occurrence of DCM and aortic dissection, wherein multivessel coronary artery dissection eventually occurred, thereby leading to advanced heart failure. She suffered from co-occurrence of DCM and aortic dissection 6 years ago. After the heart failure had briefly stabilized, the myocardial infarction due to coronary artery dissection led to worsening mitral regurgitation and decreased right ventricular function, thereby worsening the status of her heart failure. In addition to cardiovascular abnormalities, the patient was also complicated by short stature (145 cm), mild scoliosis, nonfunctioning pituitary adenoma of 1 cm in size, and retinitis pigmentosa. Coronary artery dissection is a possible complication in patients with co-occurrence of DCM and aortopathy, which could dramatically affect the clinical course of advanced heart failure.

Impact of psychiatric disorders on the hemodynamic and quality of life outcome of balloon pulmonary angioplasty in patients with chronic thromboembolic pulmonary hypertension: a retrospective study.

BACKGROUND: Balloon pulmonary angioplasty (BPA) has beneficial effects on pulmonary hemodynamics, exercise capacity, and quality of life (QOL) in patients with chronic thromboembolic pulmonary hypertension (CTEPH). Recently, emerging evidence suggests a relationship between CTEPH and psychiatric disorders (PD). However, data on the clinical efficacy of BPA in CTEPH patients with PD are lacking. METHODS: We retrospectively analyzed 75 patients with inoperable/residual CTEPH who underwent BPA and right-sided heart catheterization before the initial BPA and within 1 year after the last procedure. QOL was evaluated using the European Quality of Life Five Dimension (EQ-5D) scale in 27 patients before and after BPA sessions. Baseline and post-procedural hemodynamic, functional, and QOL parameters were compared between the patients with and without PD. RESULTS: Among the 75 participants, 22 (29.3%) patients were categorized in the PD group. Although PD group had a similar mean pulmonary artery pressure level compared with non-PD group (40 ± 7 vs. 41 ± 9 mmHg, p = 0.477), they tended to have unfavorable QOL status (0.63 ± 0.22 vs. 0.77 ± 0.19, p = 0.102). BPA significantly improved pulmonary hemodynamics, laboratory parameters and exercise tolerance in both groups. BPA also significantly improved EQ-5D scores in the non-PD group (from 0.77 ± 0.19 to 0.88 ± 0.13, p < 0.001), but the scores remained unchanged in the PD group (from 0.63 ± 0.22 to 0.67 ± 0.22, p = 0.770). During the long-term period [1,848 (1,055-2,565) days], both groups experienced similar mortality rates (PD 4.6% vs. non-PD 5.7%, p = 1.000). CONCLUSIONS: BPA improved hemodynamic and functional parameters irrespective of PD, but its effect on QOL was limited in patients with PD.

Clinical valve thrombosis and arterial embolism in a cancer patient after transcatheter aortic valve replacement.

The number of cancer patients with severe aortic stenosis and atrial fibrillation (AF) is increasing in the aging population. Transcatheter aortic valve replacement (TAVR) is an established treatment option for severe aortic stenosis with high surgical risk, including individuals with cancer. Antithrombotic therapy should be considered for post-TAVR or AF patients. However, antithrombotic management in cancer patients remains challenging due to the increased risk of both thromboembolism and bleeding. We present a case of clinical valve thrombosis and arterial embolism after transcatheter aortic valve replacement in an elderly patient with a history of metastatic pancreatic cancer and permanent atrial fibrillation under treatment of single antiplatelet therapy. Warfarin treatment after successful surgical thrombectomy to the occluded arteries improved clinical valve thrombosis, although the long-term outcome remains unclear. This case demonstrates that novel management algorithms for thromboembolism and bleeding in elderly cancer patients with AF and valvular heart disease are urgently needed.

A novel pathogenic variant located just upstream of the C-terminal Ser423-X-Ser425 phosphorylation motif in SMAD3 causing Loeys-Dietz syndrome.

OBJECTIVE: Loeys-Dietz syndrome (LDS) is a heritable disorder of connective tissue closely related to Marfan syndrome (MFS). LDS is caused by loss-of-function variants of genes that encode components of transforming growth factor-ß (TGF-ß) signaling; nevertheless, LDS type 1/2 caused by TGFBR1/2 pathogenic variants is frequently found to have paradoxical increases in TGF-ß signaling in the aneurysmal aortic wall. Here, we present a Japanese LDS family having a novel SMAD3 variant. METHODS: The proband was tested via clinical, genetic, and histological analyses. In vitro analysis was performed for pathogenic evaluation. RESULTS: The novel heterozygous missense variant of SMAD3 [c.1262G>A, p.(Cys421Tyr)], located just upstream of the C-terminal Ser423-X-Ser425 phosphorylation motif, was found in this instance of LDS type 3. This variant led to reduced phospho-SMAD3 (Ser423/Ser425) levels and transcription activity in vitro; however, a paradoxical upregulation of TGF-ß signaling was evident in the aortic wall. CONCLUSIONS: Our results revealed the presence of TGF-ß paradox in this case with the novel loss-of-function SMAD3 variant. The precise mechanism underlying the paradox is unknown, but further research is warranted to clarify the influence of the SMAD3 variant type and location on the LDS3 phenotype as well as the molecular mechanism leading to LDS3 aortopathy.

PGC-1α-mediated angiogenesis prevents pulmonary hypertension in mice.

Pulmonary hypertension (PH) is a life-threatening disease characterized by a progressive narrowing of pulmonary arterioles. Although VEGF is highly expressed in lung of patients with PH and in animal PH models, the involvement of angiogenesis remains elusive. To clarify the pathophysiological function of angiogenesis in PH, we compared the angiogenic response in hypoxia (Hx) and SU5416 (a VEGFR2 inhibitor) plus Hx (SuHx) mouse PH models using 3D imaging. The 3D imaging analysis revealed an angiogenic response in the lung of the Hx-PH, but not of the severer SuHx-PH model. Selective VEGFR2 inhibition with cabozantinib plus Hx in mice also suppressed angiogenic response and exacerbated Hx-PH to the same extent as SuHx. Expression of endothelial proliferator-activated receptor γ coactivator 1α (PGC-1α) increased along with angiogenesis in lung of Hx-PH but not SuHx mice. In pulmonary endothelial cell-specific Ppargc1a-KO mice, the Hx-induced angiogenesis was suppressed, and PH was exacerbated along with increased oxidative stress, cellular senescence, and DNA damage. By contrast, treatment with baicalin, a flavonoid enhancing PGC-1α activity in endothelial cells, ameliorated Hx-PH with increased Vegfa expression and angiogenesis. Pulmonary endothelial PGC-1α-mediated angiogenesis is essential for adaptive responses to Hx and might represent a potential therapeutic target for PH.

Effect of renal function under left ventricular assist device support on the cardiac function and clinical events after heart transplantation.

AIM: We investigated the effects of pre-transplantation renal dysfunction under left ventricular assisted device (LVAD) support on post-transplantation cardiac function, and patient prognosis after heart transplantation (HTx). METHOD: All patients who were bridged by LVAD and underwent HTx at our hospital between 2007 and 2022 were included in this study. Patients were classified into two groups based on estimated glomerular filtration rate (eGFR) before HTx: renal dysfunction (RD) group (eGFR < 60 mL/min/1.73 m2 ) and non-renal dysfunction (NRD) group. RESULT: A total of 132 patients were analyzed, of whom 48 were classified into the RD group and 84 into the NRD group (RD group, 47.9 ± 10.1 years; NRD group, 38.4 ± 11.9 years, p < .0001). Under LVAD support before HTx, the RD group tended to have a history of right ventricular failure (RD group, nine (19%); NRD group, seven (8%); p = .098). After HTx, the echocardiographic parameters did not differ between the two groups in the long term. Furthermore, more concise hemodynamic parameters, exemplified by right heart catheterization, were not significantly different between the two groups. Regarding graft rejection, no significant differences were found in acute cellular rejection and cardiac allograft vasculopathy following HTx. In contrast, patients with RD before HTx had significantly increased mortality in the chronic phase after HTx and initiation of maintenance dialysis, without any overt changes in cardiac function. CONCLUSION: Pre-transplantation renal dysfunction under LVAD support significantly affected clinical course after HTx without any overt changes in graft cardiac function.

A new assessment method for right ventricular diastolic function using right heart catheterization by pressure-volume loop.

Diastolic stiffness coefficient (ß) and end-diastolic elastance (Eed) are ventricular-specific diastolic parameters. However, the diastolic function of right ventricle had not been investigated sufficiently due to the lack of established evaluation method. We evaluated the validity of these parameters calculated using only data of right heart catheterization (RHC) and assessed it in patients with restrictive cardiomyopathy (RCM) and cardiac amyloidosis. We retrospectively analyzed 46 patients with heart failure who underwent RHC within 10 days of cardiac magnetic resonance (CMR). Right ventricular end-diastolic volume and end-systolic volume were calculated using only RHC data, which were found to be finely correlated with those obtained from CMR. ß and Eed calculated by this method were also significantly correlated with those derived from conventional method using CMR. By this method, ß and Eed were significantly higher in RCM with amyloidosis group than dilated cardiomyopathy group. In addition, the ß and Eed calculated by our method were finely correlated with E/A ratio on echocardiography. We established an easy method to estimate ß and Eed of right ventricle from only RHC. The method finely demonstrated right ventricular diastolic dysfunction in patients with RCM and amyloidosis.

Coronary Periarteritis and Pericardial Thickening Could Be Predictors for Coronary Artery Events Complicated by IgG4-Related Disease.

Background: IgG4-related disease (IgG4-RD) is a systemic disease characterized by serum IgG4 upregulation, massive infiltration of IgG4-positive plasma cells, and storiform fibrosis, which results in nodules or thickening of the involved organs. Cardiologists have recently recognized that IgG4-RD can be complicated by coronary artery events (CAEs); however, the mechanisms and clinical characteristics of this phenomenon are unknown. We evaluated the clinical signs of patients with coronary periarteritis (CP), aortic periarteritis (AP), and pericardial thickening, which are complications of IgG4-RD, to determine the contributing factors. Methods: We retrospectively examined 19 patients with IgG4-RD who attended or consulted a cardiologist in our department at the University of Tokyo Hospital between January 1, 2004, and December 31, 2021. Results: The frequency of CAEs was significantly higher in the CP group than in the non-CP group. Furthermore, the CP group had significantly lower event-free survival than the non-CP group (log-rank test, P = 0.008). However, the frequency of incidents and event-free survival for CAEs after IgG4-RD diagnosis did not differ significantly between the AP and non-AP groups. Although no statistically significant difference was present between the frequency of CAEs for those with vs without pericardial thickening, the group with pericardial thickening had significantly worse event-free survival than the group without pericardial thickening (log-rank test, P = 0.017). Conclusions: The incidence and clinical course of CAEs complicated by IgG4-RD could be predicted by identifying CP and pericardial thickening in IgG4-RD but not AP.

Contexte: La maladie liée aux immunoglobulines de type G4 (ML-IgG4) est une maladie généralisée caractérisée par une augmentation du taux sérique d'IgG4, par une infiltration massive de plasmocytes exprimant les IgG4 et par une fibrose storiforme, qui produit des nodules ou un épaississement des organes touchés. Les cardiologues ont récemment reconnu que la ML-IgG4 peut être compliquée par des événements coronariens; les mécanismes et caractéristiques cliniques de ce phénomène demeurent cependant inconnus. Nous avons évalué les signes cliniques chez des patients atteints de périartérite coronarienne (PC), de périaortite (PA) et d'épaississement du péricarde, des complications de la ML-IgG4, pour tenter d'établir les facteurs contributifs. Méthodologie: Nous avons examiné de manière rétrospective les dossiers de 19 patients atteints de ML-IgG4 qui ont été admis à notre service de l'Hôpital de l'Université de Tokyo ou qui ont consulté un cardiologue du service entre le 1er janvier 2004 et le 31 décembre 2021. Résultats: La fréquence des événements coronariens était significativement plus élevée dans le groupe PC que dans les autres groupes. Par ailleurs, le groupe PC avait une survie sans événement significativement plus courte que les autres groupes (test logarithmique par rangs; p = 0,008). En outre, la fréquence des événements coronariens et la survie sans événement coronarien après un diagnostic de ML-IgG4 ne variaient pas de manière significative entre le groupe PA et les autres groupes. Bien qu'aucune différence statistiquement significative n'ait été constatée quant à la fréquence des événements coronariens entre les patients présentant un épaississement du péricarde et les autres patients, le premier groupe affichait une survie sans événement significativement plus courte que l'autre (test logarithmique par rangs; p = 0,017). Conclusions: L'incidence et le déroulement clinique des événements coronariens compliqués par la ML-IgG4 pouvaient être anticipés dans les cas de ML-IgG4 en présence de PC et d'un épaississement du péricarde, mais pas de PA.

Novel AGL variants in a patient with glycogen storage disease type IIIb and pulmonary hypertension caused by pulmonary veno-occlusive disease: A case report.

Glycogen storage disease type III (GSD-III) is an autosomal recessive metabolic disorder caused by mutations in the AGL gene, and may develop various types of pulmonary hypertension (PH). Here, we report a case of 24-year-old man with GSD-IIIb with two novel null variants in AGL (c.2308 + 2T>C and c.3045_3048dupTACC). He developed multi-drug-resistant pulmonary veno-occlusive disease (PVOD) and was registered as a candidate for lung transplantation. No pathogenic variants were detected in previously known causative genes for pulmonary hypertension and the underlying mechanism of coincidence of two rare diseases was unknown. We discuss the association of the loss of glycogen-debranching enzyme with incident PVOD.