Transcription factor 7 like 2 promotes oligodendrocyte differentiation and remyelination

Abstract

Transcription factor 7 like 2 (TCF7L2, also termed TCF4), is a Wnt effector induced transiently in the oligodendroglial lineage. The current well accepted hypothesis is that TCF7L2 inhibits oligodendrocyte differentiation and remyelination through canonical Wnt/β‑catenin signaling. However, recent studies indicated that TCF7L2 activity is required during oligodendrocyte differentiation and remyelination. In order to clarify this, in situ hybridization, immunofluorescence and western blot analysis using in vivo TCF7L2 conditional knockout mice, were performed and it was found that TCF7L2 promotes oligodendrocyte differentiation during myelin formation and remyelination. Furthermore, it was established that TCF7L2 does not affect oligodendrocyte precursor cells during remyelination. These data are of important clinical significance to develop novel therapeutic targets to overcome multiple sclerosis and other demyelinating diseases.

Figure 1.

TCF7L2 cKO inhibits OL differentiation. (A) TCF7L2 cKO mice demonstrated that the mRNA expression level of MBP and PLP1 decreased significantly when compared with the control group in the spinal cord at P0 (n=5 per group). Magnification, ×100. (B) The TCF7L2 cKO mice demonstrated significantly decreased expression levels of MBP and PLP1 mRNA in the spinal cord when compared with the control group at P7 (n=4 per group). Magnification, ×100. (C) Quantitative analysis indicated that the mRNA expression levels of MBP and PLP1 in the TCF7L2 cKO group were significantly reduced compared with that of the control group at P0 (P<0.001). (D) Quantitative analysis further indicated that the MBP and PLP1 mRNA expression levels in the TCF7L2 cKO group were significantly lower than those of the control group at P7 (MBP, P<0.01; PLP, P<0.05). *P<0.05, **P<0.01 and ***P<0.001. Error bars indicate the standard error of the mean. TCF7L2, transcription factor 7 like 2; cKO, conditional knockout; OL, oligodendrocyte; MBP, myelin basic protein; PLP1, proteolipid protein 1. P0, the first day after birth; P7, the seventh day after birth.

Figure 2.

Expression levels of CNP and MBP proteins were reduced in the TCF7L2 cKO group. (A) Western blot analysis of the P30 corpus callosum homogenates (10 µg total protein per lane) exhibited significantly reduced expression levels of CNP (46 kDa) in the TCF7L2 cKO mice compared with the littermate controls (n=3 per group). (B) Western blot analysis indicated a significant decrease in the expression level of MBP (14, 17 and 21 kDa) in the TCF7L2 cKO mice compared with the littermate controls (n=3 per group). GAPDH (36 kDa) and β-actin (42 kDa) served as loading controls. (C) Quantitative analysis of western blot further indicated that the expression levels of CNP (46 and 48 kDa) and MBP (14, 17 and 21 kDa) were significantly lower than those of the controls. Error bars indicate standard error of the mean. **P<0.01. CNP, 2′,3′-cyclic nucleotide 3′ phosphodiesterase; MBP, myelin basic protein; TCF7L2, transcription factor 7 like 2; cKO, conditional knockout; P30, the thirtieth day after birth.

Figure 3.

Behavioral experiments from TCF7L2 cKO and littermate control mice were analyzed. The rotarod performance tests demonstrated that the riding times of the TCF7L2 cKO mice on the rotarod instrument were significantly shorter than the controls at P30, P45 and P60 (P<0.05). The riding time of the mice in the TCF7L2 cKO group decreased at P90 and P120 (P=0.131 and P=0.087, respectively). n=9 per group. Error bars indicate standard error of the mean. *P<0.05. TCF7L2, transcription factor 7 like 2; cKO, conditional knockout; CNP, 2′,3′-cyclic nucleotide 3′ phosphodiesterase; ns, not significant.

Figure 4.

TCF7L2 cKO inhibits remyelination. (A) Subsequent to feeding the mice 0.2% cuprizone for 6 weeks, myelin histochemistry with eriochrome cyanine demonstrated that there was almost no myelin in the corpus callosum samples between the two groups (n=4 per group; magnification, ×100). The bottom row presents the magnified images of the boxed areas in the top row (magnification, ×200). (B) Subsequent to returning to feeding with normal chow for 2 weeks, the corpus callosum samples were almost completely remyelinated in the control group (n=5 per group; magnification, ×100). By contrast, remyelination did not occur in the TCF7L2 cKO mice. The bottom row presents the magnified images of the boxed areas in the top row (magnification, ×200). TCF7L2, transcription factor 7 like 2; cKO, conditional knockout; CNP, 2′,3′-cyclic nucleotide 3′ phosphodiesterase.

Figure 5.

TCF7L2 promotes remyelination. After returning to feeding with normal chow for 2 weeks, immunofluorescence indicated that the expression levels of (A) APC and (B) CNP in the control mice were significantly higher than in the TCF7L2 cKO mice (n=5 per group). (C) Quantitative analysis further indicated that the expression levels of APC and CNP in the TCF7L2 cKO mice were significantly less than those of the control mice (P<0.001; n=5 per group). ***P<0.001. Error bars indicate the standard error of the mean. Magnification, ×100. TCF7L2, transcription factor 7 like 2; cKO, conditional knockout; APC, adenomatous polyposis coli; CNP, 2′,3′-cyclic nucleotide 3′ phosphodiesterase.

Figure 6.

TCF7L2 cKO does not affect OPCs during remyelination. (Aa) After returning to feeding with normal chow for 2 weeks, immunofluorescence demonstrated that the expression of PDGFRA was almost the same in the two groups (n=5 per group; magnification, ×100). (Ab) Magnified images of the boxed areas in the Aa (magnification, ×200). (B) No significant difference was demonstrated by quantitative analysis in the expression level of PDGFRA between the two groups during remyelination (P>0.05). Error bars indicate SEM. TCF7L2, transcription factor 7 like 2; cKO, conditional knockout; CNP, 2′,3′-cyclic nucleotide 3′ phosphodiesterase; PDGFRA, platelet-derived growth factor receptor α. ns, not significant.