Growth factor and cytokine gene expression in mechanically strained human osteoblast-like cells: implications for distraction osteogenesis.

OBJECTIVE: An understanding of bone cellular biology is a basic necessity to understanding events such as distraction osteogenesis. The goal of this study was to determine the effect of continuous cyclic mechanical stretch as a fundamental event in distraction osteogenesis on the expression of 3 bone growth factors, transforming growth factor-beta 1 (TGF-beta1), insulin-like growth factor 1 (IGF-1), basic fibroblast growth factor (bFGF) and 2 cytokines, interleukin (IL)-1 (IL-1) and 6 (IL-6) in human osteoblast-like cells. MATERIAL AND METHODS: A human osteoblast-like cell line, SaOS-2, capable of forming a ground substance and mineralizing it, was maintained. Cells were transferred to 6-well plates with flexible silicon bottoms grown to confluence and either subjected to tensile stretch for different time intervals or used as the control group. RNA was isolated to conduct Northern blot analysis for the expression of 3 bone growth factors, transforming TGF-beta1, IGF-1, bFGF, and 2 cytokines, IL-1 and IL-6. RESULTS: After 8 hours, mRNA for TGF-beta1 and IGF-1 increased in the experimental group, whereas bFGF decreased but cytokines IL-1 and IL-6 were not affected. At 16 hours, TGF-beta1, IGF-1, and bFGF showed increased levels of mRNA; IL-6 showed a slight increase. After 24 hours, TGF-beta1, IGF-1, bFGF, and IL-6 had increased mRNA levels. IL-1beta did never show significant alterations in mRNA production as compared with the control. CONCLUSION: Tensile stretch on osteoblast-like cells alter local regulation of bone formation, increasing the expression of bone growth factors, whereas catabolic cytokines are unaffected. These findings suggest a direct effect of mechanical strain on osteoblasts and may be the driving factors of bone growth during distraction.

The active site of the Escherichia coli MutY DNA adenine glycosylase.

Escherichia coli MutY is an adenine DNA glycosylase active on DNA substrates containing A/G, A/C, or A/8-oxoG mismatches. Although MutY can form a covalent intermediate with its DNA substrates, its possession of 3' apurinic lyase activity is controversial. To study the reaction mechanism of MutY, the conserved Asp-138 was mutated to Asn and the reactivity of this mutant MutY protein determined. The glycosylase activity was completely abolished in the D138N MutY mutant. The D138N mutant and wild-type MutY protein also possessed different DNA binding activities with various mismatches. Several lysine residues were identified in the proximity of the active site by analyzing the imino-covalent MutY-DNA intermediate. Mutation of Lys-157 and Lys-158 both individually and combined, had no effect on MutY activities but the K142A mutant protein was unable to form Schiff base intermediates with DNA substrates. However, the MutY K142A mutant could still bind DNA substrates and had adenine glycosylase activity. Surprisingly, the K142A mutant MutY, but not the wild-type enzyme, could promote a beta/delta-elimination on apurinic DNA. Our results suggest that Asp-138 acts as a general base to deprotonate either the epsilon-amine group of Lys-142 or to activate a water molecule and the resulting apurinic DNA then reacts with Lys-142 to form the Schiff base intermediate with DNA. With the K142A mutant, Asp-138 activates a water molecule to attack the C1' of the adenosine; the resulting apurinic DNA is cleaved through beta/delta-elimination without Schiff base formation.

Propofol anesthesia for outpatient oral and maxillofacial surgery.

Propofol is a sedative-hypnotic intravenous anesthetic agent that has gained wide use in outpatient oral and maxillofacial surgery since its clinical introduction in 1985. Propofol has several therapeutic advantages that make it an excellent choice for use in all phases of general anesthesia and conscious sedation. It is associated with minimal side effects, a controllable anesthetic state, and rapid recovery. This review of propofol discusses its pharmacologic character, administration, and side effects and presents anesthetic drug interaction information and comparisons.

Dr. W. Harry Archer, DDS, MA, 1905-1980 a life of dedication to oral surgery.

The life of the American oral surgeon Dr. W. Harry Archer was one that affected and inspired, and sometimes incensed, those who knew him and those who would follow him. His life long devotion to the dental specialty of oral surgery was a driving force that caused dramatic changes in aspects of not only oral surgery but of dentistry as a whole. His excellence in leadership, teaching, research, and writing were among his many accomplishments that firmly place him among the eternal giants of the dental community.

Correlation of inflammatory cytokines with arthroscopic findings in patients with temporomandibular joint internal derangements.

PURPOSE: The goal of this study was to evaluate the presence of the inflammatory cytokines interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha) within the superior temporomandibular joint (TMJ) space in patients with internal derangements and to compare these values with the pathologic findings seen arthroscopically. PATIENTS AND METHODS: Thirty patients with symptomatic TMJ dysfunction and clinical and imaging evidence of internal derangements of the TMJ were evaluated. Before entering the superior joint space with the arthroscope, 2 mL sterile saline was injected and, after 30 seconds of equilibration, was aspirated for analysis. The surgeon then performed diagnostic arthroscopy. The degree of synovitis, degeneration, percent condylar roofing, and any pathologic changes, such as perforations, were recorded. The level of total protein in each sample was ascertained, as well as the levels of IL-1 beta, IL-6, and TNF-alpha. RESULTS: Of 30 samples tested, three were discarded because of failure to gain access into the superior joint space. Of the 27 remaining samples, IL-6 showed the closest correlation with the level of acute synovitis demonstrated arthroscopically. Two of the higher IL-6 levels (167 and 324 pg/microg protein) were seen with patients with a significant disc perforation. In patients with a high degree of vascularity, IL-6 was found to be between 0 to 581 pg/microg protein with an average of 80 pg/microg protein and a median value of 43 pg/mg. These values significantly correlated with the degree of vascularity (P < or = .02). This is in comparison with the 10 remaining patients, who showed significantly fewer vascular changes arthroscopically. In these patients, the range of IL-6 was 0 to 35 pg/microg protein, with an average of 19 pg/microg protein and a median value of 14.5 pg/microg. These values significantly correlated with the smaller degree of vascularity (P < or = .02). In seven patients, the role of nonsteroidal antiinflammatory drug (NSAID) use resulted in decreased levels of IL-6, which has been noted in previous studies. In patients with higher rated redundancy of the synovial tissue, the average IL-6 level was 92 pg/microg protein, whereas the median value was 44 pg/microg protein. In patients with little or no redundant synovial tissue, an average IL-6 level of 22 pg/microg protein was present. The median value in these same joints was 15 pg/microg protein. These IL-6 values significantly correlated with the degree of redundancy (P < or = .03). The degree of degenerative change (chondromalacia, fibrillation), disc displacement (roofing), and the presence or absence of adhesions did not significantly affect the levels of IL-6 within the patients studied. The presence of IL-1 beta and TNF-alpha was not found to correlate with the arthroscopic findings in the superior joint space. CONCLUSIONS: The presence of IL-6 correlated with the degree of acute synovitis. IL-1 beta and TNF-alpha were not found in significant levels within the superior joint space. These findings correlated with those reported by other investigators. The production of IL-6 by synovial cells and its role in TMJ disease warrants further investigation.

Specific recognition of A/G and A/7,8-dihydro-8-oxoguanine (8-oxoG) mismatches by Escherichia coli MutY: removal of the C-terminal domain preferentially affects A/8-oxoG recognition.

Escherichia coli MutY is a 39 kDa adenine DNA glycosylase and 3' apurinic/apyrimidinic (AP) lyase that is active on DNA substrates containing A/G, A/C, or A/8-oxoG mismatches. 8-oxoG (7,8-dihydro-8-oxoguanine or GO) is a major stable product of oxidative damage, and A/GO mismatches may be particularly important biological substrates for MutY. Proteolytic digestion of MutY using thermolysin was found to produce two relatively stable fragments of 25 and 12 kDa. The 25 kDa fragment begins at the N terminus of MutY and spans the region homologous with E. coli endonuclease III, a DNA glycosylase/AP lyase that repairs oxidatively damaged pyrimidines. The 12 kDa fragment, which consists of much of the rest of MutY, had no detectable activity. The purified 25 kDa fragment (M25) had nearly wild-type binding and cleavage activities with A/G-mismatched substrates. Binding to A/GO-mismatched DNA, however, was dramatically reduced in M25 compared to that in intact protein. Borohydride-dependent enzyme-DNA cross-linking, which is a hallmark of the reaction of several DNA glycosylases that possess concomitant AP lyase activity, was also substantially reduced when M25 was allowed to react with A/GO-mismatched DNA. The significant differences in M25 recognition and reactivity with A/G and A/GO mismatches suggest that the C-terminal region of MutY, a region with no homologous counterpart in E. coli endonuclease III, plays an important role in the repair of mismatched DNA arising from oxidation damage.

The development of hospital dentistry in America--the first one hundred years (1850-1950).

The development of hospital dentistry in America began in the middle of the nineteenth century with the pioneering endeavors of Dr. Simon Hullihen and Dr. James Garretson who is credited with the development of the specialty of oral surgery. While starting small, the prodigious work of these and other dentists laid the foundation on which modern hospital dentistry would be built. Throughout its establishment, however, hospital dentistry would have to fight for a place in the hospital, not only with the medical community but also from within the dental community. In time though, hospital dentistry would come to have the support of the American Dental Association and the respect of the medical community.

Catalytic mechanism and DNA substrate recognition of Escherichia coli MutY protein.

Escherichia coli MutY protein cleaves A/G- or a/7,8-dihydro-8-oxo-guanine (A/GO)-containing DNA on the A-strand by N-glycosylase and apurinic/apyrimidinic endonuclease or lyase activities. In this paper, we show that MutY can be trapped in a stable covalent enzyme-DNA intermediate in the presence of sodium borohydride, a new finding that supports the grouping of MutY in that class of DNA glycosylases that possess concomitant apurinic/apyrimidinic lyase activity. To potentially help determine the substrate recognition site of MutY, mutant proteins were constructed. MutY proteins with a Gly116 --> Ala (G116A) or Asp (G116D) mutation had reduced binding affinities for both A/G- and A/GO-containing DNA substrates. The catalytic parameters, however, were differentially affected. While A/G- and A/GO-containing DNA were cleaved by MutY with specificity constants (kcat/Km) of 10 and 3.3 min-1 microM-1, respectively, MutY(G116D) cleaved these DNAs 2, 300- and 9-fold less efficiently. The catalytic activities of MutY(G116A) with A/G- and A/GO-containing DNA were about the same as that of wild-type MutY. Both MutY(G116A) and MutY(G116D) could be trapped in covalent intermediates with A/GO-containing DNA, but with lower efficiencies than the wild-type enzyme in the presence of sodium borohydride. MutY(G116A) also formed a covalent intermediate with A/G-containing DNA, but MutY(G116D) did not. Since Gly116 of MutY lies in a region that is highly conserved among several DNA glycosylases, it is likely this conserved region is in the proximity of the substrate binding and/or catalytic sites.

DNA determinants and substrate specificities of Escherichia coli MutY.

Potential DNA contacts involved in the specific interaction between the Escherichia coli MutY protein and a 40-mer oligonucleotide containing an A/G mismatch have been examined by alkylation interference techniques. Ethylation interference patterns suggest that more than five phosphates are involved in electrostatic interactions between MutY and DNA. Interestingly, MutY has more contacts on the G-strand than on the A-strand. Methylation at both the N-7 position of the mismatched G and the N-3 position of the mispaired A interfere with MutY binding. In addition to these mismatched bases, MutY also contacts purines on both sides of the mismatch. Binding and endonuclease activities of MutY were assayed with 20-mer oligonucleotides containing A/G, A/C, A/7,8-dihydro-8-oxo-guanine (A/GO), A/inosine (A/I), A/2-aminopurine (A/2AP), nebularine/G (N/G), inosine/G (I/G), 2AP/G, and 7-deaza-adenosine/G (Z/G) mispairs. The C-8 keto group of GO in A/GO contributes to a much tighter binding but weaker endonuclease activity than is seen with A/G. Because A/I is not specifically well recognized by MutY, the 2-amino group of G in A/G is essential for recognition. The C-6 keto group present in A/G but absent in A/2AP is also important for recognition. The 6-amino group of adenine appears not to be required for either binding or endonuclease activity because N/G is as good a substrate as A/G. The 2AP/G mispair is bound and cleaved weaker than is the A/G mispair. Binding and endonuclease activities are abolished when the N-7 group of A is replaced by C-7 as in the Z/G mispair. When a C-6 keto group is present as in the I/G pair, its binding by MutY is as good as for A/G, but no endonuclease activity is observed. Taken together, our data suggest that DNA sequences proximal to and specific functional groups of mismatched bases are necessary for recognition and catalysis by MutY protein.