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A Novel Role for Twist-1 in Pulp Homeostasis
K.M. Galler1,2,
A. Yasue1,
A.C. Cavender1,
P. Bialek3,
G. Karsenty4 and
R.N. D’Souza1,*
1 Department of Biomedical Sciences, Texas A&M University Health Science Center, Baylor College of Dentistry, 3302 Gaston Avenue, Dallas, TX 75246, USA;
2 Department of Restorative Dentistry and Periodontology, University of Regensburg, Germany;
3 Wyeth Research, 87 Cambridge Park Drive, Cambridge, MA, USA; and
4 Dept. of Genetics & Development, Columbia University College of Physicians and Surgeons, New York, NY, USA
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Figure 1. Phenotypic analysis of Twist-1(+/–) tissues. First maxillary molars at Day 1, von Kossa stain showing an earlier onset of organic matrix and mineral deposition (arrows). (A) = Twist-1(+/+); (B) = Twist(+/–). Hematoxylin and eosin stain of maxillary incisor (C) and alkaline phosphatase activity around blood vessels in a consecutive section from D7 Twist-1(+/–) heads (D). Signals for Dspp (E) and Col I (F) in D21 Twist-1(+/–) maxillary incisors. Dentin-like deposits within a Twist-1-deficient pulp, Masson’s trichrome, D45 first mandibular molar (G,H).
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Figure 2. RT-PCR and real-time PCR analysis of wild-type and Twist-1(+/–) animals. Total RNA was extracted and pooled from Twist-1(+/–) and Twist-1(+/+) animals of one litter at days 0, 7, and 28 for RT-PCR analysis and at D3 for quantitative real-time PCR. mRNA expression of Twist-1 in different organs. Decreased PCR product in Twist-1-deficient tissues (A). Relative gene expression of Twist-1, collagen type I (Col I), alkaline phosphatase (ALP), dentin sialophosphoprotein (Dspp), bone sialoprotein (Bsp), and osteocalcin (Oc), as observed by quantitative real-time PCR analysis in first molar organs of Twist-1(+/–) mice and wild-type littermates at post-natal D3 (B). In total, 28 molar organs/genotype/stage were used. All data are represented by mean ± standard deviation values.
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Figure 3. Cell proliferation and alkaline phosphatase activity in primary cell cultures. Data were summarized from cell cultures derived from 6 Twist-1(+/+) and 6 Twist-1(+/–) mice at post-natal D3 and D7. Cell proliferation decreased in Twist-1(+/–) cells at both stages. This effect appeared relevant, as observed during the cell culture experiment, but it is statistically insignificant (p > 0.05 for both timepoints) (A). Alkaline phosphatase activity is about two-fold increased compared with the cells derived from wild-type molar organs at post-natal D7, which is statistically highly significant (p < 0.01) (B). Student’s t test was used to analyze the differences between groups, and all data are represented by mean ± standard deviation values.
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Figure 4. Genetic rescue experiments. Whereas Twist(+/–) animals at later stages show matrix deposits and dentin-like structures within the pulpal tissues in incisors (A) and molars (B), these cannot be found in double-heterozygous animals in incisors (C) and molars (D). All sections stained with hematoxylin and eosin.
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Journal of Dental Research, Vol. 86, No. 10,
951-955 (2007)
DOI: 10.1177/154405910708601007
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