Association of serum transforming growth factor β1 with left ventricular hypertrophy in children with primary hypertension

The current study was designed to assess the association of serum transforming growth factor β1 (TGF-β1) with left ventricular hypertrophy (LVH) in children with primary hypertension. The present single-center prospective trial examined 182 patients diagnosed with primary hypertension in Children’s Hospital, Capital Institute of Pediatrics, between January 2021 and September 2022. Clinical data were analyzed, and ambulatory blood pressure was assessed for 24 h. LVH, the commonest subclinical cardiac feature of hypertension, was assessed by echocardiography. According to left ventricular geometry, cases were assigned to the LVH (n = 44) and normal geometry (n = 138) groups. Serum TGF-β1 amounts were quantitated by enzyme-linked immunosorbent assay (ELISA). Receiver operating characteristic (ROC) curves were established to analyze various variables for their predictive values in LVH. Among 182 children with primary hypertension, the concentrations of serum TGF-β1 were higher in stage 2 hypertension than in stage 1 (47.3 (38.8, 52.5) vs. 46.0 (38.6, 48.2) ng/L, Z =  − 2.376; P = 0.018). Additionally, serum TGF-β1 content showed a positive correlation with BP levels (P < 0.05). TGF-β1 amounts were significantly elevated in the LVH group compared with the normal geometry group (51.7 (46.1, 54.9) vs. 46.1 (38.7, 48.1) ng/L, Z =  − 4.324; P = 0.0000). Serum TGF-β1 content was positively associated with LVH (r = 0.321, P = 0.0000). Multivariable logistic regression analysis showed BMI (OR = 1.188, 95% CI 1.082–1.305; P = 0.0000) and elevated serum TGF-β1 content (OR = 1.063, 95% CI 1.016–1.113; P = 0.009) independently predicted LVH. A multivariable logistic regression model considering BMI and TGF-β1 content in LVH prediction was 0.771, with sensitivity and specificity of 72.7% and 70.3%, respectively. Conclusion: These data revealed an association of serum TGF-β1 with BP in children with primary hypertension. Serum TGF-β1 concentration was positively correlated with hypertensive cardiac damage. Serum TGF-β1 might constitute a valuable molecular marker for the prediction of LVH in children with primary hypertension. The combination of BMI and TGF-β1 has a certain diagnostic and predictive value for LVH in children with primary hypertension, which may provide a new reference index for early clinical identification of hypertensive cardiac damage. What is Known: • Experimental and clinical data indicated TGF-β1 is involved in BP elevation. • TGF-β1 is positively correlated with LVMI and hypertrophy in adults. What is New: • Our current study reveals an association of serum TGF-β1 with BP in children with primary hypertension. • Elevated serum TGF-β1 level is positively associated with LVH in children with primary hypertension. • The combination of BMI and TGF-β1 has a certain diagnostic and predictive value for LVH in children with primary hypertension.


Introduction
Transforming growth factor β 1 (TGF-β 1 ), a cytokine with multiple functions, belongs to the TGF-β superfamily expressed by vascular wall cells [1,2].TGF-β 1 regulates cell growth, differentiation, and growth of extracellular matrix, which is produced by myocardial cells, vascular smooth muscle cells, endothelial cells, macrophages, etc. TGF-β 1 is very important in hypertensive vascular remodeling, by affecting hemodynamics in the body, stimulating cytoendothelin production or renin-angiotensin-aldosterone system (RAAS) activation [3,4].Therefore, current studies on adults believe that TGF-β 1 is tightly associated with the pathogenesis of hypertensive target organ damage (TOD) [5].
Recently, an increasing incidence of pediatric primary hypertension has been reported.Although cardiovascular events rarely occur, subclinical TOD is common [6][7][8], and the associated damages can persist into adulthood, putting these patients at high risk of severe cardiovascular diseases [9].Lili Y et al. [10] showed that elevated blood pressure in children and adolescents was significantly correlated with high pulse wave velocity (PWV), high carotid artery intimamedia thickness (cIMT), and left ventricular hypertrophy (LVH), which was also strongly associated with cardiovascular events and all-cause mortality in adulthood.This study further supported the adverse effects of elevated blood pressure in children and adolescents on cardiovascular health in adulthood.In recent years, studies have shown that 30% of children with hypertension had LVH in the early stages [11].The evaluation of left ventricular mass (LVM) and ventricular configuration is currently considered the basis for assessing early cardiac damage in children with hypertension [12,13].As an intermediate stage in the progression of hypertensive cardiac damage from childhood to adulthood, early cardiac assessment has important values [10].Therefore, the early recognition of LVH is of great significance for the early prevention and treatment of hypertension and cardiovascular diseases in children.TGF-β 1 amounts were shown to be positively correlated with blood pressure (BP) in adult patients [14].However, previous studies have not examined serum TGF-β 1 amounts in pediatric primary hypertension cases.The current study aimed to assess the association of serum TGF-β 1 with -LVH in children with primary hypertension.

Subjects
The present single-center prospective trial enrolled 182 pediatric patients who were first diagnosed with primary hypertension hospitalized in the Children's Hospital, Capital Institute of Pediatrics (CIP; Beijing, China), between January 2021 and September 2022, including 140 males (76.9%) and 42 females (23.1%).This trial had approval from the Ethics Committee of the CIP (No: SHERLL2021021).The parents/guardians of all patients provided signed informed consent prior to enrolment.
The diagnosis was made following the indications proposed in the Chinese guidelines for the diagnosis and management of hypertension in children and adolescents [15], and all blood pressure measurements were performed using the auscultation method as recommended [16].Hypertension diagnosis was reflected by average systolic BP (SBP) and/ or diastolic BP (DBP) beyond the 95th percentiles of the auscultation measurement, after adjustments for gender, age, and height.Stages 1 and 2 hypertension were diagnosed with BP < 99th and ≥ 99th percentiles, respectively, plus 5 mmHg.
Exclusion criteria were age > 18 years; secondary hypertension associated with kidney, vascular, endocrine, or central nervous system (CNS) disease or drug treatment; and primary hypertension previously treated with antihypertensives.

Demographic and laboratory data
Demographic data include age, gender, height, weight, and body mass index (BMI) [BMI= weight (kg)/height 2 (m 2 )].The diagnostic criteria for obesity is BMI ≥ 95th percentiles, after adjustments for gender and age [17].Laboratory data include fasting blood glucose, blood lipids, plasma renin activity, angiotensin II, aldosterone, and other biochemical indicators.Biochemical indexes were quantitated in plasma or serum by routine assays.

Serum TGF-β 1 measurement
Serum human TGF-β 1 was detected by enzyme-linked immunosorbent assay (ELISA) with a specific kit (R&D Systems, USA) as directed by the manufacturer.

Blood pressure measurements
The pediatric cases were submitted to 24-h ambulatory blood pressure monitoring (ABPM) with a DMS-ABP device (DM Software, China).After training, the patients measured their BP themselves.Although daily activities were allowed, the patients were instructed to perform measurements in the resting state at 30-min and 60-min intervals in the daytime and during sleep, respectively.Nighttime was defined by recording BP in the sleep and wake periods and adjusting the values for every individual.Accordingly, 24-h SBP, 24-h DBP, daytime and nighttime SBP, and daytime and nighttime DBP were assessed.Method reliability was reflected by > 70% valid measurements [18].

Statistical analysis
SPSS 23.0 was utilized for data analysis.Two-sided P < 0.05 indicated statistical significance.Continuous data were assessed for normality by the Kolmogorov-Smirnov test.Those with skewed distribution (median and interquartile range M(Q 1 ,Q 3 )) were compared by the Mann-Whitney U-test.Categorical variates (percentage (%)) were compared by the chi-square (χ 2 ) test.The Spearman's correlation coefficient was used to assess the association between two variates.Multivariable logistic regression analysis was performed using stepwise logistic regression with forward selection and backward elimination by removing variables that had a p-value greater than 0.05.Odds ratios (ORs) with 95% confidence intervals (CIs) were determined.Receiver operating characteristic (ROC) curves were generated, and areas under the ROC curves (AUCs) were determined to assess the values of indexes in predicting LVH.

Demographics, blood pressure, and serum TGF-β 1 levels of all subjects
Totally, 182 pediatric cases of primary hypertension were included, with 140 boys (76.9%) and 42 girls (23.1%) averaging 12.7 ± 2.4 years old.Of all cases, 134 (73.6%) showed BMI values surpassing the 95th percentile of age-and gender-matched individuals and had a diagnosis of obesity.Of all 182 patients, 57 (31.3%) and 125 (68.7%) showed stages 1 and 2 hypertension, respectively.There were no significant differences between both subgroups based on age, gender, and obesity status.TGF-β 1 levels were markedly elevated in stage 2 hypertension than in stage 1 hypertension (Table 1).We analyzed the correlations between serum TGF-β 1 levels and ABPM parameters.Serum TGF-β 1 amounts had correlations with 24-h SBP and DBP, as well as daytime and nighttime SBP (P < 0.05) (Table 2).

LVH prediction in pediatric patients with primary hypertension
BMI and high serum TGF-β 1 levels independently predicted LVH (OR > 1, P < 0.05) as shown in Table 4.The predictive values of the latter indexes in pediatric LVH were determined from ROC curves.The AUCs for LVH prediction were 0.748 and 0.716 for BMI and TGF-β 1 , respectively.The cutoff values for BMI and serum TGFβ 1 were 27.7 kg/m 2 and 48.1 ng/L, respectively (Table 5).In a multivariable logistic regression model combining BMI and TGF-β 1 , an AUC of 0.771 was obtained for LVH prediction, with sensitivity and specificity of 72.7% and 70.3%, respectively (Fig. 1).

Discussion
Primary hypertension is a progressive cardiovascular disease, and vascular remodeling in hypertension continually occurs during the dynamic progression of vasoconstriction, inflammation, fibrosis, and hyperplasia [22].In the early stage of hypertension, the sympathetic nervous system and RAAS in patients are over-activated, and vasoactive substances such as adrenaline and angiotensin II have elevated levels, resulting in constant increase in the heart rate, elevated blood pressure, and accelerated remodeling of cardiovascular system [23].Meanwhile, the regulation of neurohumoral mechanism and other factors results in circadian rhythm changes in blood pressure, which are more likely to damage the patients' heart, kidney, and other target organs [24].
TGF-β 1 , a cytokine displaying multiple functions in fibrogenesis and hemodynamics [25], contributes to hypertensive vascular remodeling by affecting hemodynamics in the body, stimulating cytoendothelin production or RAAS activation [3,4].Experimental and clinical data indicated TGF-β 1 is involved in BP elevation [14,26].TGF-β 1 levels are significantly elevated in cases with hypertension, and upregulation of TGF-β 1 is associated with cardiovascular alterations [27,28].In this study, TGF-β 1 levels were markedly elevated in stage 2 hypertension compared with stage 1 hypertension, suggesting TGF-β 1 showed a positive correlation with blood pressure progression.
Children with primary hypertension have specific clinical characteristics, which can be associated with early subclinical TOD [29][30][31].Myocardial remodeling in hypertensive cases is defined as heart hypertrophy and fibrosis.LVMI and RWT represent critical indicators for evaluating early LVH [12].LVH often constitutes the major sign of early heart injury in pediatric hypertensive cases.Representing an intermediary phase of cardiac damage between childhood and adulthood, early cardiac damage is critical for disease evaluation [32,33].In this study, of all 182 patients, 24.2% had LVH, indicating that some children with primary hypertension already had early cardiac damage at presentation.
Previous studies have shown that the etiological mechanism of LVH is complex, involving many factors such as intracellular and external stimuli, genes, and heredity.
It was found that TGF-β 1 is positively correlated with LVMI and hypertrophy in adults [34].TGF-β 1 is the cytokine most potently regulating LVH and promoting fibrogenesis.Multiple reports have demonstrated a close association of serum TGF-β 1 with LVH [35,36].TGF-β 1 is produced in cardiomyocytes and myocardial fibrocytes, as one of the most important cytokines regulating myocardial hypertrophy.First, TGF-β 1 is the initial factor in the synthesis and deposition of collagen fibers and other extracellular components, which stimulates the synthesis of new contractile proteins in the myocardium, thereby inducing the re-expression of embryonic genes.Second, TGF-β 1 regulates the synthesis of extracellular matrix proteins and increases the production of collagen, proteoglycan, and fibronectin, which blunts matrix degradation by decreasing collagenase synthesis and upregulating protease inhibitors.
As shown above, elevated TGF-β 1 was associated with LVH.Serum TGF-β 1 levels were elevated in LVH cases

Limitation
Limitations of the study lie in that, firstly, this was a smallsample single-center study, which requires multicenter trials for validation.Secondly, children with normal blood pressure were not included as the control group.Thirdly, because of no follow-up, the long-term effects of TGF-β 1 in children with primary hypertension could not be examined.The above cases should be further examined for an extended period of time, particularly monitoring TGF-β 1 amounts, which may help clearly reveal the exact effects of TGF-β 1 on BP and LVH.

Conclusion
In conclusion, the current data revealed an association of serum TGF-β 1 with BP in children with primary hypertension.Serum TGF-β 1 content was positively correlated with hypertensive cardiac damage.Serum TGF-β 1 might represent an important molecular marker for LVH prediction in children with primary hypertension.The combination of BMI and TGF-β 1 has a certain diagnostic and predictive value in LVH in children with primary hypertension, which may provide a new reference index for early clinical identification of hypertensive cardiac damage.

Table 1
Demographics and serum TGF-β 1 levels of all subjects

Table 2
Serum TGF-β 1 levels and ABPM parameters Bold values represent P values less than 0.05 TGF-β 1 transforming growth factor β 1 , SBP systolic blood pressure, DBP diastolic blood pressure

Table 3
Bold values represent P values less than 0.05 BMI body mass index, TGF-β 1 transforming growth factor β 1 , CI confidence interval Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made.The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material.If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder.To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.