Learning more about the role of Latent transforming growth factor-β binding protein–1 (LTBP–1) – a knockout approach in mice

Aims: The latent transforming growth factor-β binding proteins (LTBP) are a family of widely expressed multidomain glycoproteins. cDNA for four different LTBP (LTBP–1, –2, –3, –4) was cloned from different sources and also from different species. The LTBP genes set up a subfamily of the extracellular fibrillin proteins, which constitute the backbone of extracellular filaments. We focused our work on LTBP–1 which is expressed in different organs [1], forms intracellular complexes with transforming growth factor-β (TGF-β1, –2, –3), target the growth factor to the extracellular matrix, and is a key player in controlling wound healing. Methods: We generated a mouse model system, in which both isoforms of the LTBP–1 gene, the short (LTBP–1S) and the long isoform (LTBP–1L), were deleted. We evaluated in detail liver sections from wild type and knock out littermates by Sirius Red staining as well as the measurement of serum parameters (AST, ALT, bilirubin) relevant for liver function. In addition, RT-PCR analysis was performed to monitor the expression level of the other LTBP isoforms (LTBP–2, –3, –4) in cultured hepatic stellate cells (HSC) and skin fibroblasts. Comparative gene expression profiles from HSC were generated using PIQOR™ Immunology Microarrays (Miltenyi Biotec). Results: Delicate differences between wild type and knockout animals were detectable in bone morphology. Generally, the head was more compact in LTBP–1 deficient mice than in normal controls. In cultured HSC and skin fibroblasts, the relative transcript levels of other LTBP isoforms were not altered as proven by RT-PCR. Microarray analysis substantiated the crucial influence of LTBP–1 on TGF-β-dependent genes and lead to the identification of novel gene targets controlled by the LTBP–1/TGF-β network. Conclusions: The staining of liver tissues from LTBP–1 nulls displayed, that the absence of LTBP–1 does not induce any signs of hepatic dysfunction. However, our microarray data analysis clearly demonstrates that LTBP–1-deficiency is strongly associated with differences in expressing and controlling of various TGF-β-dependent genes. Therefore, we conclude that the LTBP–1 deficiency will be critical in situations when TGF-β signalling is pronounced, i.e. fibrogenesis.