2004). Because this protein forms a latent complex with TGF-��, one of the most profibrogenic molecules known to date, it would be likely that the fibrosis present though in the liver of AhR-null mice would also show an increase in LTBP-1 protein expression. Therefore, we analysed LTBP-1 protein expression in the same areas of the liver where a high content in total collagen, ��-actin and vimentin were detected. The results obtained supported this hypothesis, as AhR?/? livers had a marked increase in LTBP-1 protein expression, particularly in the areas surrounding the periportal space (Fig. 1i,j). LTBP-1 content was also increased in the portal fibrotic areas of AhR-null mice. However, this protein could be either deposited or synthesized in situ in the fibrotic compartment.
To answer this question, we detected the anatomical distribution of LTBP-1 mRNA in liver sections using in situ hybridization (Fig. 2). LTBP-1 antisense probe (LTBP-AS) detected increased levels of mRNA expression in the portal areas of AhR?/? as compared to wild-type mice (Fig. 2a,b). Immunohistochemistry for LTBP-1 in an equivalent area of the liver revealed overexpression of LTBP-1 protein (Fig. 2c). In situ hybridization for LTBP-1 was specific, as the addition of the sense LTBP-1 probe (LTBP-1-SS) to either AhR?/? or AhR+/+ liver sections did not produce any signal. Figure 2 In situ hybridization of latent TGF-��-binding protein-1 (LTBP-1) mRNA. Serial 4-��m frozen sections containing a typical portal triad from wild-type AhR (AhR+/+) and null AhR (AhR?/?) mice livers were hybridized with LTBP-1 .
.. It has been reported that disruption of AhR gene expression produced an elevation in the basal levels of TGF-�� protein in the liver (Zaher et al. 1998). According to this, TGF-�� protein in AhR?/? livers was markedly expressed around the portal areas (Fig. 3a,b). Additionally, as LTBP-1 and TGF-�� are known to form a latent complex, these two mRNAs would likely colocalize to the cells in the fibrotic area. However, when we compared the anatomic distribution of both TGF-�� and LTBP-1 mRNAs using slides from the same portal fibrotic region, we found that TGF-�� mRNA was expressed in all hepatic parenchyma, whereas LTBP-1 was mainly present in the portal area. Therefore, the pattern for mRNA expression of both genes was not restricted to the same location in the liver (Fig. 3a�Cc).
Moreover, Fig. 3 also shows that TGF-�� mRNA levels in AhR?/? were very similar to those found in AhR+/+ liver. The TGF-�� antisense probe used (TGF-��1-AS) was specific, because the addition of TGF-�� sense probe (TGF-��1-SS) to equivalent liver tissue sections did not produce any signal (Fig. 3f,g). Figure 3 Immunostaining for TGF-�� (panels A, B) of wild-type Dacomitinib AhR (AhR+/+) and null AhR (AhR?/?) mice livers. In situ hybridization of TGF-�� mRNA. Serial 4-��m frozen sections containing a typical portal triad from wild-type …
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