Transforming growth factor-beta (TGF- β) signaling in paravertebral muscles in juvenile and adolescent idiopathic scoliosis

Abstract

Most researchers agree that idiopathic scoliosis (IS) is a multifactorial disease influenced by complex genetic and environmental factors. The onset of the spinal deformity that determines the natural course of the disease, usually occurs in the juvenile or adolescent period. Transforming growth factors β (TGF-βs) and their receptors, TGFBRs, may be considered as candidate genes related to IS susceptibility and natural history. This study explores the transcriptional profile of TGF-βs, TGFBRs, and TGF-β responsive genes in the paravertebral muscles of patients with juvenile and adolescent idiopathic scoliosis (JIS and AIS, resp.). Muscle specimens were harvested intraoperatively and grouped according to the side of the curve and the age of scoliosis onset. The results of microarray and qRT-PCR analysis confirmed significantly higher transcript abundances of TGF-β2, TGF-β3, and TGFBR2 in samples from the curve concavity of AIS patients, suggesting a difference in TGF-β signaling in the pathogenesis of juvenile and adolescent curves. Analysis of TGF-β responsive genes in the transcriptomes of patients with AIS suggested overrepresentation of the genes localized in the extracellular region of curve concavity: LTBP3, LTBP4, ITGB4, and ITGB5. This finding suggests the extracellular region of paravertebral muscles as an interesting target for future molecular research into AIS pathogenesis.

Figure 1

Heat map of fluorescence signal intensities of TGFB 1–3 and TGFBRs 1–3. Heat map of fluorescence signals intensities of 10 mRNA IDs of genes encoding TGF-β 1–3 and their receptors TGFBR 1–3 in paravertebral muscles of curve concavity (M1) and curve convexity (M2) in a group of juvenile (A) and adolescent idiopathic scoliosis (B).

Figure 2

Evaluation of the transcriptional profile of TGF-βs and their receptors TGFBRs in paravertebral muscles of JIS group A and AIS group B by qRT-PCR. Box and whisker plots illustrating statistically significant (nonparametric U Mann-Whitney test, P < 0.05) results of the comparison of the mRNA abundance of TGF-βs and TGFBRs between concave and convex sides of the curve in both analyzed groups. AM1, AM2, respectively, paravertebral muscle of concave and convex side of the curve in JIS group and BM1, BM2 samples from concave and convex side of the curve in AIS group.

Figure 3

TGF-β responsive genes in the paravertebral muscles of JIS and AIS patients. Matrix plot illustrating the degree of differentiation of 1050 mRNA IDs of TGF-β responsive genes between the transcriptomes of muscular tissue in dependence of the side of the curve (M1 versus M2) and the age of scoliosis onset (A versus B). Red spots upregulated genes, blue spots downregulated genes. M1, M2: respectively muscular tissue samples from curve concavity and convexity, A: juvenile idiopathic scoliosis; B: adolescent idiopathic scoliosis.

Figure 4

Venn diagram of two way ANOVA test for TGF-β responsive genes. Venn diagram illustrating the number of TGF-β responsive genes with a P value < 0.05 (two way ANOVA) differentially expressed between the sides of the curve (M1 versus M2), age of scoliosis onset (A versus B) and their interactions.

Figure 5

Overrepresentation test of upregulated genes differentiating muscular transcriptomes in JIS and AIS—GO molecular function. Overlaid area chart of difference presenting the results of an overrepresentation test of GO molecular function of upregulated genes differentiating concave and convex paravertebral muscle transcriptomes in JIS and AIS group. ∗ above the peak mark statistically significant result of overrepresentation test, P < 0.05 calculated with Bonferroni multiple correction.

Figure 6

Overrepresentation test of upregulated genes differentiating muscular transcriptomes in AIS—GO cellular component localization. Bar chart of difference presenting the results of an overrepresentation test of GO cellular component localization of upregulated genes differentiating concave and convex paravertebral muscle transcriptomes in the AIS group. Yellow bars indicate statistically significant results of overrepresentation test, P < 0.05 calculated with Bonferroni multiple correction.