Orofacial Granulomatosis among Pediatric Patients Well Controlled by Corticosteroid Treatment: A Rare Case Series.

Orofacial granulomatosis (OFG) is a rare disease entity characterized by nonnecrotizing granulomatous inflammation in the oral and maxillofacial regions, typically characterized by recurrent or persistent edema, primarily in the lips and occasionally in the gingiva. OFG is often associated with Crohn's disease and sarcoidosis, and an accurate diagnosis requires systemic examination of patients. Pediatric patients possess unique oral conditions where dental plaque rapidly forms, especially during tooth replacement due to tooth crowding. Moreover, controlling oral hygiene can be challenging, rendering it difficult to distinguish plaque-induced gingivitis from nonplaque-induced gingivitis. We elucidate the reports of pediatric patients who developed OFG in the lips and/or gingiva alone, which was well controlled through corticosteroid treatment. The patients demonstrated recurrent lips and/or gingival swelling with redness, which failed to improve despite oral health care and treatment with antibiotics and/or corticosteroid ointment. Incision biopsy was performed, which demonstrated granulomatous inflammation. Further systemic examination ruled out Crohn's disease and sarcoidosis and confirmed OFG diagnosis. Corticosteroid treatment orally or through gargling was administered to the patients, which provided improvement of symptoms after 1 month. As OFG may be associated with intractable diseases, monitoring the patient regularly is crucial. Pediatric patients with OFG require a collaborative approach with pediatricians and pediatric dentists to manage their oral and overall health.

Three-dimensional structure of nylon hydrolase and mechanism of nylon-6 hydrolysis.

We performed x-ray crystallographic analyses of the 6-aminohexanoate oligomer hydrolase (NylC) from Agromyces sp. at 2.0 Å-resolution. This enzyme is a member of the N-terminal nucleophile hydrolase superfamily that is responsible for the degradation of the nylon-6 industry byproduct. We observed four identical heterodimers (27 kDa + 9 kDa), which resulted from the autoprocessing of the precursor protein (36 kDa) and which constitute the doughnut-shaped quaternary structure. The catalytic residue of NylC was identified as the N-terminal Thr-267 of the 9-kDa subunit. Furthermore, each heterodimer is folded into a single domain, generating a stacked αßßα core structure. Amino acid mutations at subunit interfaces of the tetramer were observed to drastically alter the thermostability of the protein. In particular, four mutations (D122G/H130Y/D36A/E263Q) of wild-type NylC from Arthrobacter sp. (plasmid pOAD2-encoding enzyme), with a heat denaturation temperature of T(m) = 52 °C, enhanced the protein thermostability by 36 °C (T(m) = 88 °C), whereas a single mutation (G111S or L137A) decreased the stability by ∼10 °C. We examined the enzymatic hydrolysis of nylon-6 by the thermostable NylC mutant. Argon cluster secondary ion mass spectrometry analyses of the reaction products revealed that the major peak of nylon-6 (m/z 10,000-25,000) shifted to a smaller range, producing a new peak corresponding to m/z 1500-3000 after the enzyme treatment at 60 °C. In addition, smaller fragments in the soluble fraction were successively hydrolyzed to dimers and monomers. Based on these data, we propose that NylC should be designated as nylon hydrolase (or nylonase). Three potential uses of NylC for industrial and environmental applications are also discussed.

X-ray crystallographic and mutational analysis of the NylC precursor: catalytic mechanism of autocleavage and substrate hydrolysis of nylon hydrolase.

Nylon hydrolase (NylC), a member of the N-terminal nucleophile (Ntn) hydrolase superfamily, is responsible for the degradation of various aliphatic nylons, including nylon-6 and nylon-66. NylC is initially expressed as an inactive precursor (36 kDa), but the precursor is autocatalytically cleaved at Asn266/Thr267 to generate an active enzyme composed of 27 and 9 kDa subunits. We isolated various mutants with amino acid changes at the catalytic centre. X-ray crystallographic analysis revealed that the NylC precursor forms a doughnut-shaped quaternary structure composed of four monomers (molecules A-D) with D2 symmetry. Catalytic residues in the precursor are covered by loop regions at the A/B interface (equivalent to the C/D interface). However, the catalytic residues are exposed to the solvent environment through autocleavage followed by movements of the loop regions. T267A, D306A and D308A mutations resulted in a complete loss of autocleavage. By contrast, in the T267S mutant, autocleavage proceeded slowly at a constant reaction rate (k = 2.8 × 10-5  s-1 ) until complete conversion, but the reaction was inhibited by K189A and N219A mutations. Based on the crystallographic and molecular dynamic simulation analyses, we concluded that the Asp308-Asp306-Thr267 triad, resembling the Glu-Ser-Ser triad conserved in Ntn-hydrolase family enzymes, is responsible for autocleavage and that hydrogen-bonding networks connecting Thr267 with Lys189 and Asn219 are required for increasing the nucleophilicity of Thr267-OH in both the water accessible and water inaccessible systems. Furthermore, we determined that NylC employs the Asp308-Asp306-Thr267 triad as catalytic residues for substrate hydrolysis, but the reaction requires Lys189 and Tyr146 as additional catalytic/substrate-binding residues specific for nylon hydrolysis.

Elevated expression of calcineurin subunits during active mineralization of developing mouse molar teeth.

Calcineurin is a Ca(2+) /calmodulin-dependent protein phosphatase consisting of two subunits - catalytic subunit A (CnA) and regulatory subunit B (CnB) - and plays a critical role in transducing Ca(2+) signals into cellular responses. In this study, we investigated the expression of calcineurin in the mouse developing tooth. In-situ hybridization detected mRNAs for the CnAα and CnAß isoforms of CnA and for the CnB1 isoform of CnB in the upper molar tooth germ at embryonic day 15. Immunohistochemistry with antibodies specific for CnAα, CnAß, and CnB1 showed strong immunoreactivity of these proteins in secretory-stage ameloblasts and in odontoblasts during dentin formation. CnAß and CnB1 were strongly immunoreactive in ruffle-ended ameloblasts at the enamel-maturation stage. In ameloblasts and odontoblasts, we also noted different subcellular distributions of CnAα and CnAß. From these data, temporal profiles of calcineurin expression appear to correlate with active mineralization in tooth development. Furthermore, the distinct subcellular distribution of the two CnA subunits may reflect their distinct substrates or responsive sites within single cells, thus contributing to the diversity of calcineurin-dependent cellular responses during active tooth mineralization.

Spontaneously formed semipermeable organic-inorganic hybrid vesicles permitting molecular weight selective transmembrane passage.

Semipermeable organic-inorganic hybrid vesicles coated with a siloxane surface were spontaneously formed by simple dispersion of an organoalkoxysilane lipid in water. The hybrid vesicles allow the permeation of hydrophilic small molecules across the membrane without an introduction of a pore-forming protein.