Hypomaturation Enamel Defects in Klk4 Knockout/LacZ Knockin Mice

Kallikrein 4 (Klk4) is believed to play an essential role in enamel biomineralization, because defects in KLK4 cause hypomaturation amelogenesis imperfecta. We used gene targeting to generate a knockin mouse that replaces the Klk4 gene sequence, starting at the translation initiation site, with a lacZ reporter gene. Correct targeting of the transgene was confirmed by Southern blot and PCR analyses. Histochemical X-gal (5-bromo-4-chloro-3-indolyl-β-d-galactopyranoside) staining demonstrated expression of β-galactosidase in maturation stage ameloblasts. No X-gal staining was observed in secretory stage ameloblasts or in odontoblasts. Retained enamel proteins were observed in the maturation stage enamel of the Klk4 null mouse, but not in the Klk4 heterozygous or wild-type mice. The enamel layer in the Klk4 null mouse was normal in thickness and contained decussating enamel rods but was rapidly abraded following weaning, despite the mice being maintained on soft chow. In function the enamel readily fractured within the initial rod and interrod enamel above the parallel enamel covering the dentino-enamel junction. Despite the lack of Klk4 and the retention of enamel proteins, significant levels of crystal maturation occurred (although delayed), and the enamel achieved a mineral density in some places greater than that detected in bone and dentin. An important finding was that individual enamel crystallites of erupted teeth failed to grow together, interlock, and function as a unit. Instead, individual crystallites seemed to spill out of the enamel when fractured. These results demonstrate that Klk4 is essential for the removal of enamel proteins and the proper maturation of enamel crystals. Kallikrein 4 (Klk4) is believed to play an essential role in enamel biomineralization, because defects in KLK4 cause hypomaturation amelogenesis imperfecta. We used gene targeting to generate a knockin mouse that replaces the Klk4 gene sequence, starting at the translation initiation site, with a lacZ reporter gene. Correct targeting of the transgene was confirmed by Southern blot and PCR analyses. Histochemical X-gal (5-bromo-4-chloro-3-indolyl-β-d-galactopyranoside) staining demonstrated expression of β-galactosidase in maturation stage ameloblasts. No X-gal staining was observed in secretory stage ameloblasts or in odontoblasts. Retained enamel proteins were observed in the maturation stage enamel of the Klk4 null mouse, but not in the Klk4 heterozygous or wild-type mice. The enamel layer in the Klk4 null mouse was normal in thickness and contained decussating enamel rods but was rapidly abraded following weaning, despite the mice being maintained on soft chow. In function the enamel readily fractured within the initial rod and interrod enamel above the parallel enamel covering the dentino-enamel junction. Despite the lack of Klk4 and the retention of enamel proteins, significant levels of crystal maturation occurred (although delayed), and the enamel achieved a mineral density in some places greater than that detected in bone and dentin. An important finding was that individual enamel crystallites of erupted teeth failed to grow together, interlock, and function as a unit. Instead, individual crystallites seemed to spill out of the enamel when fractured. These results demonstrate that Klk4 is essential for the removal of enamel proteins and the proper maturation of enamel crystals. Dental enamel is composed of highly ordered, very long crystals of calcium hydroxyapatite (Ca10(PO4)6(OH)2). Mature enamel crystallites are about 70 nm wide and 30 nm thick, but are of unmeasurable length (1.Warshawsky H. Nanci A. J. Dent. Res. Spec. No. 1982; : 1504-1514Google Scholar), probably extending all the way from the dentin layer to the surface of the tooth (2.Daculsi G. Menanteau J. Kerebel L.M. Mitre D. Calcif. Tissue Int. 1984; 36: 550-555Crossref PubMed Scopus (122) Google Scholar). Enamel crystallites are organized into bundles called rods, with about 10,000 parallel crystals in a rod (3.Daculsi G. Kerebel B. J. Ultrastruct. Res. 1978; 65: 163-172Crossref PubMed Scopus (129) Google Scholar). Each enamel rod is the product of a single ameloblast, the cell type that forms a continuous sheet over the developing enamel and orchestrates its formation. Dental enamel of erupted teeth is ∼95% mineral (by weight) (4.Deakins M. Volker J.F. J. Dent. Res. 1941; 20: 117-121Crossref Scopus (30) Google Scholar), with most of the non-mineral component being water. Protein comprises <1% of its weight. Forming enamel, however, is over 30% protein (5.Fukae M. Shimizu M. Arch. Oral Biol. 1974; 19: 381-386Crossref PubMed Scopus (74) Google Scholar). Much of the protein is reabsorbed by ameloblasts and degraded in lysosomes (6.Sasaki T. Histochemistry. 1984; 80: 263-268Crossref PubMed Scopus (28) Google Scholar, 7.Smid J.R. Young W.G. Monsour P.A. Eur. J. Oral Sci. 2001; 109: 260-266Crossref PubMed Scopus (3) Google Scholar), but extracellular proteases also play a role in matrix protein removal (8.Smith C.E. Pompura J.R. Borenstein S. Fazel A. Nanci A. Anat. Rec. 1989; 224: 292-316Crossref PubMed Scopus (88) Google Scholar, 9.Bartlett J.D. Simmer J.P. Crit. Rev. Oral Biol. Med. 1999; 10: 425-441Crossref PubMed Scopus (225) Google Scholar, 10.Simmer J.P. Hu J.C. Connect. Tissue Res. 2002; 43: 441-449Crossref PubMed Scopus (0) Google Scholar).Dental enamel formation is divided into secretory, transition, and maturation stages (11.Reith E.J. J. Ultrastruct. Res. 1970; 30: 111-151Crossref PubMed Scopus (182) Google Scholar, J. Ultrastruct. Res. PubMed Scopus Google Scholar). the secretory enamel crystals grow in the crystals the enamel layer Enamel crystallites a at the secretory surface of the cell mineral rapidly on the and very on (3.Daculsi G. Kerebel B. J. Ultrastruct. Res. 1978; 65: 163-172Crossref PubMed Scopus (129) Google Scholar, J. Biol. PubMed Scopus Google Scholar, S. M. T. Calcif. Tissue Int. PubMed Scopus (88) Google Scholar). the of the secretory stage the enamel crystals are and the enamel layer as a is as as but about of the mineral as when the tooth C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). the secretory stage is a the ameloblasts of enamel proteins T. M. Simmer J.P. H. T. Biol. 1999; PubMed Scopus Google and to maturation ameloblasts Tissue 1974; PubMed Scopus Google Scholar). the maturation mineral is on the of enamel crystallites B. G. Kerebel L.M. J. Dent. Res. PubMed Google Scholar), grow in and thickness is by with crystals Oral and Scholar, of Oral and Scholar). maturation the protein by from 30 to (5.Fukae M. Shimizu M. Arch. Oral Biol. 1974; 19: 381-386Crossref PubMed Scopus (74) Google Scholar), and of the enamel mineral is The of mineral is in the of significant protein and the crystals to grow and to C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google secretory stage enamel proteins are M. T. Sci. 80: PubMed Scopus Google Scholar, M. Res. PubMed Scopus Google Scholar), T. J. Res. PubMed Scopus Google Scholar, J. Biol. PubMed Scopus Google Scholar, M. T. T. Shimizu M. Simmer J.P. J. Dent. Res. PubMed Scopus Google Scholar), and M. T. T. Shimizu M. Simmer J.P. J. Dent. Res. PubMed Scopus Google Scholar, H. Simmer J.P. J. Dent. Res. PubMed Scopus Google Scholar). These proteins function enamel and the by mice are to the developing teeth and enamel B. G. T. T. S. G. J. Biol. 2001; PubMed Scopus Google Scholar, S. T. B. T. G. Nanci A. J. Biol. PubMed Scopus Google Scholar, J.C. Hu C.E. Simmer J.P. J. Biol. PubMed Scopus (129) Google Scholar). These are or are to in as or that to developing teeth M. Biol. PubMed Scopus Google Scholar, A. J. Biol. PubMed Scopus Google Scholar). the enamel extracellular matrix proteins are for enamel are not of the enamel to tooth enamel proteins are by proteases and reabsorbed by ameloblasts. extracellular matrix proteases are in the of enamel matrix used used J.C. A. J. of Scopus Google and 4 (Klk4) J.P. A. J. of is with and by secretory stage ameloblasts J.D. Simmer J.P. J. PubMed Scopus Google Scholar, J.D. Eur. J. Oral Sci. PubMed Scopus Google Scholar, J.C. S. J.D. Simmer J.P. Eur. J. Oral Sci. 2002; PubMed Scopus Google Scholar). for the of observed in secretory stage enamel proteins Hu J.D. Simmer J.P. J. Dent. Res. 1999; PubMed Scopus Google and to the by ameloblasts the secretory mice enamel that is and than enamel rod and to the surface J. Simmer J.P. H. J.D. J. Biol. 2002; PubMed Scopus Google Scholar, J.D. C.E. J. Dent. Res. PubMed Scopus Google Scholar). the secretory stage enamel proteins, expression to to developing teeth A. Simmer J.P. J.D. PubMed Scopus Google Scholar), as is the when the gene is Simmer J.P. J.D. Hu J.C. J. Med. PubMed Scopus Google Scholar, D. M. J. Dent. Res. PubMed Scopus Google Scholar, Hu Simmer J.P. J.D. Hu J.C. J. Dent. Res. PubMed Scopus Google is a that is by and maturation stage ameloblasts but is not by secretory stage ameloblasts J.P. M. T. T. J. Shimizu M. Hu J.D. J. Dent. Res. PubMed Scopus Google Scholar, J.C. T. H. J.D. Simmer J.P. J. Dent. Res. PubMed Scopus Google Scholar). Klk4 also by the that dentin M. T. T. H. M. Simmer J.P. S. J. Dent. Res. 2002; PubMed Scopus Google Scholar). Klk4 Hu J.C. J.D. Simmer J.P. Eur. J. Oral Sci. 2002; PubMed Scopus Google Scholar, M. D. PubMed Scopus Google and is of proteases M. 2001; PubMed Scopus Google Scholar, J. B. G. Biol. PubMed Scopus Google Scholar, H. G. J. A. M. J. Biol. PubMed Scopus (129) Google and M. H. J.D. J. Biol. PubMed Scopus Google Scholar, Res. PubMed Scopus Google Scholar). most proteins by Klk4 is in most the Sci. 1999; PubMed Scopus Google and PubMed Scopus Google Scholar). Much on the role of Klk4 in Klk4 is in A. T. J. Res. PubMed Scopus Google Scholar), in M. 2002; PubMed Scopus Google Scholar, A. A. 2002; PubMed Scopus Google Scholar, A. H. B. Res. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar), and A. S. J. H. Res. 2001; Google Scholar, A. D. M. S. Res. 2001; Google Scholar, J. B. B. PubMed Scopus Google Scholar, A. M. M. A. Biol. PubMed Scopus Google Scholar). Despite on the role of Klk4 in very is about the normal expression and function of Klk4 in of function in KLK4 a hypomaturation enamel in the of defects in the D. S. J. Med. PubMed Scopus Google into the role of Klk4 in normal enamel and to the normal and of Klk4 used gene targeting to out normal Klk4 the Klk4 with the gene β-galactosidase reporter in mice. We demonstrate that Klk4 is not by secretory stage but is by ameloblasts in enamel formation and is for the proper removal of enamel proteins, the of enamel and for of the enamel Klk4 knockin mouse that Klk4 is not by secretory stage ameloblasts. Klk4 expression in is with in the of Klk4 expression by ameloblasts is at the of the The enamel layer in the Klk4 null mouse normal thickness and is composed of decussating enamel No Klk4 expression by was These to the that Klk4 not function the secretory stage of amelogenesis and that the observed enamel defects in the Klk4 mice are to the lack of and of Klk4 in the and maturation of the stages of amelogenesis is the in and thickness of highly enamel crystals. is a that the of ameloblasts and These about C.E. Nanci A. Dent. Res. PubMed Scopus Google Scholar). is is the of the is an in the of calcium and into the matrix with mineral and a in matrix S. T. M. Arch. Oral Biol. 36: PubMed Scopus Google Scholar, C.E. M. Dent. Res. 10: PubMed Scopus Google Scholar). The of the ameloblasts are to that to in the of degraded enamel In maturation are with and the maturation stage when or enamel proteins C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). The is with the by the of by S. T. Tissue Res. PubMed Scopus Google and its into the matrix by J.F. S. Biol. PubMed Scopus Google in with the J. Dent. Res. PubMed Scopus Google maturation of enamel crystals the removal of enamel proteins from the is believed that enamel proteins to the of enamel at or and the of on the crystal surface J.P. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). enamel crystals are in of calcium crystal is observed the enamel is with or to protein J. Connect. Tissue Res. 1989; Google Scholar). enamel crystal in J. Res. PubMed Scopus Google Scholar), and the of for calcium hydroxyapatite is by with and Klk4 D. J. J. Dent. Res. PubMed Scopus Google Scholar). Despite in and that the of for proper enamel maturation of of that as as a of enamel maturation is in the maturation stage to the removal of enamel proteins C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). enamel crystallites not in the Klk4 null mouse, in and thickness the maturation stage despite the retention of enamel The of calcium and enamel proteins from the as proteins from a hydroxyapatite however, a in enamel maturation in the Klk4 heterozygous and null mice the in the the enamel an density in the of the mouse is not in the heterozygous mouse, because the enamel to a for in the enamel of the Klk4 null mouse to the of the crystals to that the individual crystallites grow into with interlock, and as a unit. of the maturation by the of enamel proteins, the The removal of enamel proteins, to the for the for crystals to within the of the secretory stage enamel M. T. S. T. J. Dent. Res. PubMed Scopus (28) Google Scholar). The enamel of the Klk4 null mouse is with the that the enamel protein the removal of the is a that and enamel crystals by the The role of Klk4 is to the of the matrix its removal (by and into to for the of enamel to the of a role for Klk4 in enamel formation was the of a in a for an in KLK4 of a with hypomaturation amelogenesis D. S. J. Med. PubMed Scopus Google Scholar). We a Klk4 null mouse that a and of its on the extracellular but is that Klk4 in to the of maturation stage ameloblasts the of of the was observed in the Klk4 null Klk4 also in or in as The Klk4 null mouse a for of the role of Klk4 in normal and Dental enamel is composed of highly ordered, very long crystals of calcium hydroxyapatite (Ca10(PO4)6(OH)2). Mature enamel crystallites are about 70 nm wide and 30 nm thick, but are of unmeasurable length (1.Warshawsky H. Nanci A. J. Dent. Res. Spec. No. 1982; : 1504-1514Google Scholar), probably extending all the way from the dentin layer to the surface of the tooth (2.Daculsi G. Menanteau J. Kerebel L.M. Mitre D. Calcif. Tissue Int. 1984; 36: 550-555Crossref PubMed Scopus (122) Google Scholar). Enamel crystallites are organized into bundles called rods, with about 10,000 parallel crystals in a rod (3.Daculsi G. Kerebel B. J. Ultrastruct. Res. 1978; 65: 163-172Crossref PubMed Scopus (129) Google Scholar). Each enamel rod is the product of a single ameloblast, the cell type that forms a continuous sheet over the developing enamel and orchestrates its formation. Dental enamel of erupted teeth is ∼95% mineral (by weight) (4.Deakins M. Volker J.F. J. Dent. Res. 1941; 20: 117-121Crossref Scopus (30) Google Scholar), with most of the non-mineral component being water. Protein comprises <1% of its weight. Forming enamel, however, is over 30% protein (5.Fukae M. Shimizu M. Arch. Oral Biol. 1974; 19: 381-386Crossref PubMed Scopus (74) Google Scholar). Much of the protein is reabsorbed by ameloblasts and degraded in lysosomes (6.Sasaki T. Histochemistry. 1984; 80: 263-268Crossref PubMed Scopus (28) Google Scholar, 7.Smid J.R. Young W.G. Monsour P.A. Eur. J. Oral Sci. 2001; 109: 260-266Crossref PubMed Scopus (3) Google Scholar), but extracellular proteases also play a role in matrix protein removal (8.Smith C.E. Pompura J.R. Borenstein S. Fazel A. Nanci A. Anat. Rec. 1989; 224: 292-316Crossref PubMed Scopus (88) Google Scholar, 9.Bartlett J.D. Simmer J.P. Crit. Rev. Oral Biol. Med. 1999; 10: 425-441Crossref PubMed Scopus (225) Google Scholar, 10.Simmer J.P. Hu J.C. Connect. Tissue Res. 2002; 43: 441-449Crossref PubMed Scopus (0) Google Scholar). Dental enamel formation is divided into secretory, transition, and maturation stages (11.Reith E.J. J. Ultrastruct. Res. 1970; 30: 111-151Crossref PubMed Scopus (182) Google Scholar, J. Ultrastruct. Res. PubMed Scopus Google Scholar). the secretory enamel crystals grow in the crystals the enamel layer Enamel crystallites a at the secretory surface of the cell mineral rapidly on the and very on (3.Daculsi G. Kerebel B. J. Ultrastruct. Res. 1978; 65: 163-172Crossref PubMed Scopus (129) Google Scholar, J. Biol. PubMed Scopus Google Scholar, S. M. T. Calcif. Tissue Int. PubMed Scopus (88) Google Scholar). the of the secretory stage the enamel crystals are and the enamel layer as a is as as but about of the mineral as when the tooth C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). the secretory stage is a the ameloblasts of enamel proteins T. M. Simmer J.P. H. T. Biol. 1999; PubMed Scopus Google and to maturation ameloblasts Tissue 1974; PubMed Scopus Google Scholar). the maturation mineral is on the of enamel crystallites B. G. Kerebel L.M. J. Dent. Res. PubMed Google Scholar), grow in and thickness is by with crystals Oral and Scholar, of Oral and Scholar). maturation the protein by from 30 to (5.Fukae M. Shimizu M. Arch. Oral Biol. 1974; 19: 381-386Crossref PubMed Scopus (74) Google Scholar), and of the enamel mineral is The of mineral is in the of significant protein and the crystals to grow and to C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). The secretory stage enamel proteins are M. T. Sci. 80: PubMed Scopus Google Scholar, M. Res. PubMed Scopus Google Scholar), T. J. Res. PubMed Scopus Google Scholar, J. Biol. PubMed Scopus Google Scholar, M. T. T. Shimizu M. Simmer J.P. J. Dent. Res. PubMed Scopus Google Scholar), and M. T. T. Shimizu M. Simmer J.P. J. Dent. Res. PubMed Scopus Google Scholar, H. Simmer J.P. J. Dent. Res. PubMed Scopus Google Scholar). These proteins function enamel and the by mice are to the developing teeth and enamel B. G. T. T. S. G. J. Biol. 2001; PubMed Scopus Google Scholar, S. T. B. T. G. Nanci A. J. Biol. PubMed Scopus Google Scholar, J.C. Hu C.E. Simmer J.P. J. Biol. PubMed Scopus (129) Google Scholar). These are or are to in as or that to developing teeth M. Biol. PubMed Scopus Google Scholar, A. J. Biol. PubMed Scopus Google Scholar). the enamel extracellular matrix proteins are for enamel are not of the enamel to tooth enamel proteins are by proteases and reabsorbed by ameloblasts. extracellular matrix proteases are in the of enamel matrix used used J.C. A. J. of Scopus Google and 4 (Klk4) J.P. A. J. of Scholar). matrix 4 dentino-enamel knockin matrix 4 dentino-enamel knockin is with and by secretory stage ameloblasts J.D. Simmer J.P. J. PubMed Scopus Google Scholar, J.D. Eur. J. Oral Sci. PubMed Scopus Google Scholar, J.C. S. J.D. Simmer J.P. Eur. J. Oral Sci. 2002; PubMed Scopus Google Scholar). for the of observed in secretory stage enamel proteins Hu J.D. Simmer J.P. J. Dent. Res. 1999; PubMed Scopus Google and to the by ameloblasts the secretory mice enamel that is and than enamel rod and to the surface J. Simmer J.P. H. J.D. J. Biol. 2002; PubMed Scopus Google Scholar, J.D. C.E. J. Dent. Res. PubMed Scopus Google Scholar). the secretory stage enamel proteins, expression to to developing teeth A. Simmer J.P. J.D. PubMed Scopus Google Scholar), as is the when the gene is Simmer J.P. J.D. Hu J.C. J. Med. PubMed Scopus Google Scholar, D. M. J. Dent. Res. PubMed Scopus Google Scholar, Hu Simmer J.P. J.D. Hu J.C. J. Dent. Res. PubMed Scopus Google Scholar). Klk4 is a that is by and maturation stage ameloblasts but is not by secretory stage ameloblasts J.P. M. T. T. J. Shimizu M. Hu J.D. J. Dent. Res. PubMed Scopus Google Scholar, J.C. T. H. J.D. Simmer J.P. J. Dent. Res. PubMed Scopus Google Scholar). Klk4 also by the that dentin M. T. T. H. M. Simmer J.P. S. J. Dent. Res. 2002; PubMed Scopus Google Scholar). Klk4 Hu J.C. J.D. Simmer J.P. Eur. J. Oral Sci. 2002; PubMed Scopus Google Scholar, M. D. PubMed Scopus Google and is of proteases M. 2001; PubMed Scopus Google Scholar, J. B. G. Biol. PubMed Scopus Google Scholar, H. G. J. A. M. J. Biol. PubMed Scopus (129) Google and M. H. J.D. J. Biol. PubMed Scopus Google Scholar, Res. PubMed Scopus Google Scholar). most proteins by Klk4 is in most the Sci. 1999; PubMed Scopus Google and PubMed Scopus Google Scholar). Much on the role of Klk4 in Klk4 is in A. T. J. Res. PubMed Scopus Google Scholar), in M. 2002; PubMed Scopus Google Scholar, A. A. 2002; PubMed Scopus Google Scholar, A. H. B. Res. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar), and A. S. J. H. Res. 2001; Google Scholar, A. D. M. S. Res. 2001; Google Scholar, J. B. B. PubMed Scopus Google Scholar, A. M. M. A. Biol. PubMed Scopus Google Scholar). Despite on the role of Klk4 in very is about the normal expression and function of Klk4 in of function in KLK4 a hypomaturation enamel in the of defects in the D. S. J. Med. PubMed Scopus Google Scholar). into the role of Klk4 in normal enamel and to the normal and of Klk4 used gene targeting to out normal Klk4 the Klk4 with the gene β-galactosidase reporter in mice. We demonstrate that Klk4 is not by secretory stage but is by ameloblasts in enamel formation and is for the proper removal of enamel proteins, the of enamel and for of the enamel Klk4 knockin mouse that Klk4 is not by secretory stage ameloblasts. Klk4 expression in is with in the of Klk4 expression by ameloblasts is at the of the The enamel layer in the Klk4 null mouse normal thickness and is composed of decussating enamel No Klk4 expression by was These to the that Klk4 not function the secretory stage of amelogenesis and that the observed enamel defects in the Klk4 mice are to the lack of and of Klk4 in the and maturation of the stages of amelogenesis is the in and thickness of highly enamel crystals. is a that the of ameloblasts and These about C.E. Nanci A. Dent. Res. PubMed Scopus Google Scholar). is is the of the is an in the of calcium and into the matrix with mineral and a in matrix S. T. M. Arch. Oral Biol. 36: PubMed Scopus Google Scholar, C.E. M. Dent. Res. 10: PubMed Scopus Google Scholar). The of the ameloblasts are to that to in the of degraded enamel In maturation are with and the maturation stage when or enamel proteins C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). The is with the by the of by S. T. Tissue Res. PubMed Scopus Google and its into the matrix by J.F. S. Biol. PubMed Scopus Google in with the J. Dent. Res. PubMed Scopus Google maturation of enamel crystals the removal of enamel proteins from the is believed that enamel proteins to the of enamel at or and the of on the crystal surface J.P. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). enamel crystals are in of calcium crystal is observed the enamel is with or to protein J. Connect. Tissue Res. 1989; Google Scholar). enamel crystal in J. Res. PubMed Scopus Google Scholar), and the of for calcium hydroxyapatite is by with and Klk4 D. J. J. Dent. Res. PubMed Scopus Google Scholar). Despite in and that the of for proper enamel maturation of of that as as a of enamel maturation is in the maturation stage to the removal of enamel proteins C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). enamel crystallites not in the Klk4 null mouse, in and thickness the maturation stage despite the retention of enamel The of calcium and enamel proteins from the as proteins from a hydroxyapatite however, a in enamel maturation in the Klk4 heterozygous and null mice the in the the enamel an density in the of the mouse is not in the heterozygous mouse, because the enamel to a for in the enamel of the Klk4 null mouse to the of the crystals to that the individual crystallites grow into with interlock, and as a unit. of the maturation by the of enamel proteins, the The removal of enamel proteins, to the for the for crystals to within the of the secretory stage enamel M. T. S. T. J. Dent. Res. PubMed Scopus (28) Google Scholar). The enamel of the Klk4 null mouse is with the that the enamel protein the removal of the is a that and enamel crystals by the The role of Klk4 is to the of the matrix its removal (by and into to for the of enamel to the of a role for Klk4 in enamel formation was the of a in a for an in KLK4 of a with hypomaturation amelogenesis D. S. J. Med. PubMed Scopus Google Scholar). We a Klk4 null mouse that a and of its on the extracellular but is that Klk4 in to the of maturation stage ameloblasts the of of the was observed in the Klk4 null Klk4 also in or in as The Klk4 null mouse a for of the role of Klk4 in normal and The Klk4 knockin mouse that Klk4 is not by secretory stage ameloblasts. Klk4 expression in is with in the of Klk4 expression by ameloblasts is at the of the The enamel layer in the Klk4 null mouse normal thickness and is composed of decussating enamel No Klk4 expression by was These to the that Klk4 not function the secretory stage of amelogenesis and that the observed enamel defects in the Klk4 mice are to the lack of and of Klk4 in the and maturation The of the stages of amelogenesis is the in and thickness of highly enamel crystals. is a that the of ameloblasts and These about C.E. Nanci A. Dent. Res. PubMed Scopus Google Scholar). is is the of the is an in the of calcium and into the matrix with mineral and a in matrix S. T. M. Arch. Oral Biol. 36: PubMed Scopus Google Scholar, C.E. M. Dent. Res. 10: PubMed Scopus Google Scholar). The of the ameloblasts are to that to in the of degraded enamel In maturation are with and the maturation stage when or enamel proteins C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). The is with the by the of by S. T. Tissue Res. PubMed Scopus Google and its into the matrix by J.F. S. Biol. PubMed Scopus Google in with the J. Dent. Res. PubMed Scopus Google Scholar). The maturation of enamel crystals the removal of enamel proteins from the is believed that enamel proteins to the of enamel at or and the of on the crystal surface J.P. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). enamel crystals are in of calcium crystal is observed the enamel is with or to protein J. Connect. Tissue Res. 1989; Google Scholar). enamel crystal in J. Res. PubMed Scopus Google Scholar), and the of for calcium hydroxyapatite is by with and Klk4 D. J. J. Dent. Res. PubMed Scopus Google Scholar). Despite in and that the of for proper enamel maturation of of that as as a of enamel maturation is in the maturation stage to the removal of enamel proteins C.E. Crit. Rev. Oral Biol. Med. PubMed Scopus Google Scholar). enamel crystallites not in the Klk4 null mouse, in and thickness the maturation stage despite the retention of enamel The of calcium and enamel proteins from the as proteins from a hydroxyapatite however, a in enamel maturation in the Klk4 heterozygous and null mice the in the the enamel an density in the of the mouse is not in the heterozygous mouse, because the enamel to a for The in the enamel of the Klk4 null mouse to the of the crystals to that the individual crystallites grow into with interlock, and as a unit. of the maturation by the of enamel proteins, the The removal of enamel proteins, to the for the for crystals to within the of the secretory stage enamel M. T. S. T. J. Dent. Res. PubMed Scopus (28) Google Scholar). The enamel of the Klk4 null mouse is with the that the enamel protein the removal of the is a that and enamel crystals by the The role of Klk4 is to the of the matrix its removal (by and into to for the of enamel crystals. to the of a role for Klk4 in enamel formation was the of a in a for an in KLK4 of a with hypomaturation amelogenesis D. S. J. Med. PubMed Scopus Google Scholar). We a Klk4 null mouse that a and of its on the extracellular but is that Klk4 in to the of maturation stage ameloblasts the of of the was observed in the Klk4 null Klk4 also in or in as The Klk4 null mouse a for of the role of Klk4 in normal and We of the of and for and in M. of the of for and in of the of and of the of for and to gene and the of at the Klk4 knockin and the mouse