Nucleus-specific translation and assembly of acetylcholinesterase in multinucleated muscle cells.

Multinucleated skeletal muscle fibers synthesize cell surface and secreted oligomeric forms of acetylcholinesterase (AChE) that accumulate at specialized locations on the cell surface, such as sites of nerve-muscle contact. Using allelic variants of the AChE polypeptide chains as genetic markers, we show that nuclei homozygous for either the alpha or beta alleles residing in chimeric myotubes preferentially translate their AChE mRNAs on their respective ERs. These results indicate that the events of transcription, translation, and assembly of this membrane protein are compartmentalized into nuclear domains in multinucleated cells, and provide the structural basis for the possible localized expression and regulation of synaptic components at the neuromuscular junctions of vertebrate skeletal muscle fibers.

Cell surface acetylcholinesterase molecules on multinucleated myotubes are clustered over the nucleus of origin.

Multinucleated skeletal muscle fibers are compartmentalized with respect to the expression and organization of several intracellular and cell surface proteins including acetylcholinesterase (AChE). Mosaic muscle fibers formed from homozygous myoblasts expressing two allelic variants of AChE preferentially translate and assemble the polypeptides in the vicinity of the nucleus encoding the mRNA (Rotundo, R. L. 1990. J. Cell Biol. 110:715-719). To determine whether the locally synthesized AChE molecules are targeted to specific regions of the myotube surface, primary quail myoblasts were mixed with mononucleated cells of the mouse muscle C2/C12 cell line and allowed to fuse, forming heterospecific mosaic myotubes. Cell surface enzyme was localized by immunofluorescence using an avian AChE-specific monoclonal antibody. HOECHST 33342 was used to distinguish between quail and mouse nuclei in myotubes. Over 80% of the quail nuclei exhibited clusters of cell surface AChE in mosaic quail-mouse myotubes, whereas only 4% of the mouse nuclei had adjacent quail AChE-positive regions of membrane, all of which were located next to a quail nucleus. In contrast, membrane proteins such as Na+/K+ ATPase, which are not restricted to specific regions of the myotube surface, are free to diffuse over the entire length of the fiber. These studies indicate that the AChE molecules expressed in multinucleated muscle fibers are preferentially transported and localized to regions of surface membrane overlying the nucleus of origin. This targeting could play an important role in establishing and maintaining specialized cell surface domains such as the neuromuscular and myotendinous junctions.

Transplantation of quail collagen-tailed acetylcholinesterase molecules onto the frog neuromuscular synapse.

The highly organized pattern of acetylcholinesterase (AChE) molecules attached to the basal lamina of the neuromuscular junction (NMJ) suggests the existence of specific binding sites for their precise localization. To test this hypothesis we immunoaffinity purified quail globular and collagen-tailed AChE forms and determined their ability to attach to frog NMJs which had been pretreated with high-salt detergent buffers. The NMJs were visualized by labeling acetylcholine receptors (AChRs) with TRITC-alpha-bungarotoxin and AChE by indirect immunofluorescence; there was excellent correspondence (>97%) between the distribution of frog AChRs and AChE. Binding of the exogenous quail AChE was determined using a species-specific monoclonal antibody. When frog neuromuscular junctions were incubated with the globular G4/G2 quail AChE forms, there was no detectable binding above background levels, whereas when similar preparations were incubated with the collagen-tailed A12 AChE form >80% of the frog synaptic sites were also immunolabeled for quail AChE attached. Binding of the A12 quail AChE was blocked by heparin, yet could not be removed with high salt buffer containing detergent once attached. Similar results were obtained using empty myofiber basal lamina sheaths produced by mechanical or freeze-thaw damage. These experiments show that specific binding sites exist for collagen-tailed AChE molecules on the synaptic basal lamina of the vertebrate NMJ and suggest that these binding sites comprise a "molecular parking lot" in which the AChE molecules can be released, retained, and turned over.

The dystroglycan complex is necessary for stabilization of acetylcholine receptor clusters at neuromuscular junctions and formation of the synaptic basement membrane.

The dystrophin-associated protein (DAP) complex spans the sarcolemmal membrane linking the cytoskeleton to the basement membrane surrounding each myofiber. Defects in the DAP complex have been linked previously to a variety of muscular dystrophies. Other evidence points to a role for the DAP complex in formation of nerve-muscle synapses. We show that myotubes differentiated from dystroglycan-/- embryonic stem cells are responsive to agrin, but produce acetylcholine receptor (AChR) clusters which are two to three times larger in area, about half as dense, and significantly less stable than those on dystroglycan+/+ myotubes. AChRs at neuromuscular junctions are similarly affected in dystroglycan-deficient chimeric mice and there is a coordinate increase in nerve terminal size at these junctions. In culture and in vivo the absence of dystroglycan disrupts the localization to AChR clusters of laminin, perlecan, and acetylcholinesterase (AChE), but not rapsyn or agrin. Treatment of myotubes in culture with laminin induces AChR clusters on dystroglycan+/+, but not -/- myotubes. These results suggest that dystroglycan is essential for the assembly of a synaptic basement membrane, most notably by localizing AChE through its binding to perlecan. In addition, they suggest that dystroglycan functions in the organization and stabilization of AChR clusters, which appear to be mediated through its binding of laminin.

Acetylcholinesterase clustering at the neuromuscular junction involves perlecan and dystroglycan.

Formation of the synaptic basal lamina at vertebrate neuromuscular junction involves the accumulation of numerous specialized extracellular matrix molecules including a specific form of acetylcholinesterase (AChE), the collagenic-tailed form. The mechanisms responsible for its localization at sites of nerve- muscle contact are not well understood. To understand synaptic AChE localization, we synthesized a fluorescent conjugate of fasciculin 2, a snake alpha-neurotoxin that tightly binds to the catalytic subunit. Prelabeling AChE on the surface of Xenopus muscle cells revealed that preexisting AChE molecules could be recruited to form clusters that colocalize with acetylcholine receptors at sites of nerve-muscle contact. Likewise, purified avian AChE with collagen-like tail, when transplanted to Xenopus muscle cells before the addition of nerves, also accumulated at sites of nerve-muscle contact. Using exogenous avian AChE as a marker, we show that the collagenic-tailed form of the enzyme binds to the heparan-sulfate proteoglycan perlecan, which in turn binds to the dystroglycan complex through alpha-dystroglycan. Therefore, the dystroglycan-perlecan complex serves as a cell surface acceptor for AChE, enabling it to be clustered at the synapse by lateral migration within the plane of the membrane. A similar mechanism may underlie the initial formation of all specialized basal lamina interposed between other cell types.

Absence of alpha-syntrophin leads to structurally aberrant neuromuscular synapses deficient in utrophin.

The syntrophins are a family of structurally related proteins that contain multiple protein interaction motifs. Syntrophins associate directly with dystrophin, the product of the Duchenne muscular dystrophy locus, and its homologues. We have generated alpha-syntrophin null mice by targeted gene disruption to test the function of this association. The alpha-Syn(-/)- mice show no evidence of myopathy, despite reduced levels of alpha-dystrobrevin-2. Neuronal nitric oxide synthase, a component of the dystrophin protein complex, is absent from the sarcolemma of the alpha-Syn(-/)- mice, even where other syntrophin isoforms are present. alpha-Syn(-/)- neuromuscular junctions have undetectable levels of postsynaptic utrophin and reduced levels of acetylcholine receptor and acetylcholinesterase. The mutant junctions have shallow nerve gutters, abnormal distributions of acetylcholine receptors, and postjunctional folds that are generally less organized and have fewer openings to the synaptic cleft than controls. Thus, alpha-syntrophin has an important role in synapse formation and in the organization of utrophin, acetylcholine receptor, and acetylcholinesterase at the neuromuscular synapse.

Periodontal conditions of sites treated with gingival-augmentation surgery compared to untreated contralateral homologous sites: a 10- to 27-year long-term study.

BACKGROUND: The aim of this retrospective long-term split-mouth study was to compare the periodontal conditions of sites treated with gingival-augmentation procedures to untreated homologous contralateral sites over a long period of time (10 to 27 years). METHODS: Fifty-five subjects with 73 sites (test group) lacking attached gingiva associated with recessions were treated by means of submarginal free gingival grafts (SMFGGs) and marginal free gingival grafts (MFGGs). The 73 contralateral homologous sites (control group), with or without recession and with or without attached gingiva, were not treated. Patients were recalled every 4 months during the follow-up period (10 to 27 years). Clinical variables, including recession depth, amount of keratinized tissue (KT), and probing depth (PD), were measured in treated and untreated sites at baseline, at 1 year, and at the end of the follow-up period. RESULTS: At the end of the follow-up period, recession was reduced in all treated sites (1.5 +/- 1.0 mm for SMFGG and 1.3 +/- 0.9 mm for MFGG), whereas it was increased in the untreated sites (-0.7 +/- 0.7 mm for SMFGG and -1.0 +/- 0.5 mm for MFGG). In the treated sites, the increased KT remained quite stable during the follow-up period. PD remained stable (1 mm) in the treated and untreated sites. CONCLUSIONS: The sites treated with gingival-augmentation surgery showed a tendency for coronal displacement of the gingival margin with a reduction in recession. The contralateral untreated sites showed a tendency for apical displacement of the gingival margin with an increase in the existing recessions.

Compartmentalization of acetylcholinesterase mRNA and enzyme at the vertebrate neuromuscular junction.

Acetylcholinesterase (AChE) is concentrated at the vertebrate neuromuscular synapse. To determine whether increased transcript levels could underlie this selective accumulation, we employed a quantitative reverse transcription polymerase chain reaction-based assay to determine mRNA copy number in samples as small as single neuromuscular junctions (NMJs) and a microassay to measure AChE enzyme activity at single synapses. Our results show that AChE mRNA is an intermediate transcript at NMJs, whereas in noninnervated regions of muscle fibers, AChE transcripts are either undetectable or rare. In contrast, alpha-actin transcript levels in the same samples are similar in junctional and extrajunctional regions. However, compared with AChE enzyme activity and alpha-actin mRNA levels, the levels of AChE transcripts at NMJs are highly variable. These results indicate that AChE mRNA and protein expression are compartmentalized at the vertebrate NMJ and provide a direct approach toward dissecting the molecular events leading from synaptic activation to plastic changes in gene expression at single vertebrate synapses.

Assembly and regulation of acetylcholinesterase at the vertebrate neuromuscular junction.

The collagen-tailed form of acetylcholinesterase (ColQ-AChE) is the major if not unique form of the enzyme associated with the neuromuscular junction (NMJ). This enzyme form consists of catalytic and non-catalytic subunits encoded by separate genes, assembled as three enzymatic tetramers attached to the three-stranded collagen-like tail (ColQ). This synaptic form of the enzyme is tightly attached to the basal lamina associated with the glycosaminoglycan perlecan. Fasciculin-2 is a snake toxin that binds tightly to AChE. Localization of junctional AChE on frozen sections of muscle with fluorescent Fasciculin-2 shows that the labeled toxin dissociates with a half-life of about 36 h. The fluorescent toxin can subsequently be taken up by the muscle fibers by endocytosis giving the appearance of enzyme recycling. Newly synthesized AChE molecules undergo a lengthy series of processing events before final transport to the cell surface and association with the synaptic basal lamina. Following co-translational glycosylation the catalytic subunit polypeptide chain interacts with several molecular chaperones, glycosidases and glycosyltransferases to produce a catalytically active enzyme that can subsequently bind to one of two non-catalytic subunits. These molecular chaperones can be rate limiting steps in the assembly process. Treatment of muscle cells with a synthetic peptide containing the PRAD attachment sequence and a KDEL retention signal results in a large increase in assembled and exportable AChE, providing an additional level of post-translational control. Finally, we have found that Pumilio2, a member of the PUF family of RNA-binding proteins, is highly concentrated at the vertebrate neuromuscular junction where it plays an important role in regulating AChE translation through binding to a highly conserved NANOS response element in the 3'-UTR. Together, these studies define several new levels of AChE regulation in electrically excitable cells.

Secretion of gamma-glutamyl hydrolase in vitro.

gamma-Glutamyl hydrolase (also known as conjugase) is a ubiquitous enzyme that has the capacity to cleave folyl- and antifolylpolyglutamates. This study has revealed that the enzyme is secreted by primary cultures of rat hepatocytes and by H35 hepatoma cells. H35 cells have lower cellular levels of gamma-glutamyl hydrolase than do hepatocytes but secrete a greater proportion of gamma-glutamyl hydrolase. More than 99% of the total enzyme from H35 cells accumulated in the medium after 48 h. The cells were shown to remain intact during the secretion period since lactate dehydrogenase, dihydrofolate reductase, and lysosomal hydrolases other than gamma-glutamyl hydrolase were retained within the cell. Using the substrate 4-amino-10-methyl-pteroyldiglutamate (4-NH2-10-CH3-Pte-Glu2), the intracellular and secreted enzyme form(s) from H35 cells were found to have the following properties (a) Km values of 24.3 +/- 3.7 microM and 34.8 +/- 8.6 microM, respectively, and (b) maximal activity at pH 5 to 7 and apparent molecular weights of 120,000 by gel filtration. Both the cellular and secreted enzymes convert 4-NH2-10-CH3-PteGlu4 and pteroylpentaglutamate acid, to the corresponding monoglutamates with little or no appearance of intermediate chain length polyglutamates. This suggests that both act primarily as endopeptidases. Thus far, the cellular and secreted enzymes cannot be differentiated although the current studies do not establish this point unequivocally. Alterations in the cellular and secreted H35 cell gamma-glutamyl hydrolase levels in response to changes in culture conditions revealed that glutamine enhances activity while insulin diminishes it. Other transformed cells found to secrete this protein are Hep-G2 human hepatoma, JAR human choriocarcinoma, HeLa, and rat glioma. gamma-Glutamyl hydrolase could not be detected in medium conditioned by human MCF-7 breast cancer cells, and relatively low activities were found in the medium from CCRF-CEM or K562 leukemia cells. These studies directly establish for the first time the secretion of gamma-glutamyl hydrolase in vitro.

Local control of acetylcholinesterase gene expression in multinucleated skeletal muscle fibers: individual nuclei respond to signals from the overlying plasma membrane.

Nuclei in multinucleated skeletal muscle fibers are capable of expressing different sets of muscle-specific genes depending on their locations within the fiber. Here we test the hypothesis that each nucleus can behave autonomously and responds to signals generated locally on the plasma membrane. We used acetylcholinesterase (AChE) as a marker because its transcripts and protein are concentrated at the neuromuscular and myotendenous junctions. First, we show that tetrodotoxin (TTX) reversibly suppresses accumulation of cell surface AChE clusters, whereas veratridine or scorpion venom (ScVn) increase them. AChE mRNA levels are also regulated by membrane depolarization. We then designed chambered cultures that allow application of sodium channel agonists or antagonists to restricted regions of the myotube surface. When a segment of myotube is exposed to TTX, AChE cluster formation is suppressed only on that region. Conversely, ScVn increases AChE cluster formation only where in contact with the muscle surface. Likewise, both the synthesis and secretion of AChE are shown to be locally regulated. Moreover, using in situ hybridization, we show that the perinuclear accumulation of AChE transcripts also depends on signals that each nucleus receives locally. Thus AChE can be up- and downregulated in adjacent regions of the same myotubes. These results indicate that individual nuclei are responding to locally generated signals for cues regulating gene expression.

Acetylcholinesterase biosynthesis and transport in tissue culture.

The enzyme acetylcholinesterase consists of a family of molecular forms differing in subunit composition, solubility properties, and subcellular location. The use of a variety of reversible or irreversible active site-directed ligands with different membrane permeability properties permits the selective inactivation of separate pools of enzyme molecules. The application of these inhibitors together with standard biochemical techniques has permitted a detailed characterization of the synthesis and metabolism of the secretory and membrane-bound acetylcholinesterase in tissue-cultured cells. These techniques, with minor modifications and appropriate controls, can also be applied to the study of AChE metabolism in organ culture and in vivo.

Targeting acetylcholinesterase molecules to the neuromuscular synapse.

The functional integrity of the neuromuscular synapse requires that sufficient numbers of acetylcholinesterase (AChE) molecules be localized on the specialized extracellular matrix between the nerve terminal and the post-synaptic membrane. Multiple interrelated levels of regulation are necessary to accomplish this complex task including the spatial and temporal restriction of AChE mRNA expression within the muscle fiber, local translation and assembly of AChE polypeptides, and focused accumulation of AChE molecules on the extracellular matrix. This is accomplished in part through the organization of other extracellular matrix molecules into a complex which further associates with acetylcholine receptors and their accompanying molecules. Finally, the mature neuromuscular junction contains molecules which can act as receptors for the attachment of AChE which in turn may allow for the turnover of this enzyme at the synapse. This brief review will focus mainly on contributions from our laboratory towards understanding the mechanisms involved in organizing AChE molecules at the neuromuscular synapse.

The characteristics and consequences of folate depletion in hepatoma cells in vitro by inhibition of dihydrofolate reductase.

Growth of rat hepatoma cells in subtoxic concentrations of the DHFR inhibitor metoprine caused a marked time and concentration dependent reduction in cellular folates. As much as 75% total cellular folates can be lost without impairing growth. Increasing the concentration of metoprine into a range that causes inhibition of growth results in no further reduction in cellular folates. This effect is presumably mediated through inhibition of DHFR and several mechanisms are discussed which may account for these results. Cells grown in medium in which the concentration of folate is changed from 4 microM to 20 nM had intracellular folate levels that were reduced 85%. This is nearly the same reduction caused by treating cells grown in normal medium (4 microM folate) with continuous, subtoxic levels of metoprine. The reduction in cellular folates caused by growth in nM folic acid caused enhanced growth inhibitory activity of several antifolates. On a concentration basis metoprine was 12-fold more active under these conditions, PDDF was 37-fold more active and DDATHF was 44-fold more active. The reason for the enhanced sensitivity to PDDF and DDATHF may also be analogous to the reason for their synergism with the low concentration of metoprine and trimetrexate (12).

The properties and function of gamma-glutamyl hydrolase and poly-gamma-glutamate.

gamma-Glutamyl hydrolase is a ubiquitous enzyme that has the capacity to cleave gamma-glutamyl bonds of cellular folyl- and antifolylpoly-gamma-glutamates. This study has revealed that the enzyme is secreted by primary cultures of rat hepatocytes and by H35 hepatoma cells. It was found that more than 99% of the total enzyme from H35 cells accumulated in the medium after 48 hr incubation with the serum-free medium. The cells were shown to remain intact during the secretion period since lactate dehydrogenase, dihydrofolate reductase and lysosomal hydrolases other than gamma-glutamyl hydrolase were retained within the cell. When PteGlu5 (folylGlu4) is used as a substrate the initial product is PteGlu (folate), and there is no appearance of intermediate chain length pteroyl polyglutamates. Therefore, the secreted and cellular gamma-glutamyl hydrolase from hepatoma cells appears to be an endopeptidase. Polyclonal antibodies to the poly-gamma-glutamate substrates of the enzyme were prepared and characterized. The antibodies recognize the structural differences between alpha- and gamma-glutamyl linkages but appear equally active with PteGlu5 and its analogs such as 4-NH2-10-CH3PteGlu5 and pABAGlu5. The affinity of the antibodies is related to the gamma-glutamyl structure since L-glutamic acid, folate or p-aminobenzoic acid are inactive with the antibodies. Furthermore, poly-gamma-glutamate has lower affinity for the antibodies than the poly-gamma-glutamate derivatives of PteGlu, 4-NH2-10-CH3PteGlu or pABA.

Viral etiology of gingival recession. A case report.

Herpes simplex virus-type I (HSV-1) is responsible for both primary and recurrent infections of the oral mucosa. The aim of this case report is to show how HSV-1 may cause periodontal damage such as gingival recession. A 26-year-old male patient presented in a private office for the treatment of gingival recessions. He reported that the recessions had appeared suddenly with marginal inflammation of the gingiva and vesicle formation; within a few hours, the gingival tissue had been completely destroyed. The lesions were accompanied by pain, fever, and regional lymphadenopathy. Two weeks later, the patient returned complaining of a recurrence accompanied by pain and lymphadenopathy. The following day, the patient's condition had worsened and the depth of the recession had increased. A biopsy was taken for histological examination. A free epithelial-connective tissue graft was performed. Histological and direct immunofluorescence examinations confirmed the herpetic origin of the lesion. Eight months after surgery, a new herpetic lesion was detected in correspondence to the gingival margin of the first lower right premolar; therefore, acyclovir was prescribed. After 1 week, the antiviral therapy was completely successful; the gingival lesion disappeared, and no recession of the soft tissue margin was observed. Based on these clinical features, diagnosis of gingival recession induced by HSV-1 must be carried out at an early stage to establish a successful therapy.

Oral Acanthosis nigricans as a marker of internal malignancy. A case report.

BACKGROUND: Acanthosis nigricans (AN) is a rare mucocutaneous condition that can involve the oral tissues. There are 2 clinical forms of AN: benign and malignant. Benign AN is related to systemic diseases such as diabetes and obesity or can be induced by drugs such as systemic corticosteroids, nicotinic acid, estrogens, insulin, and fusidic acid. Malignant AN appears in association with tumors such as lung, ovarian, breast, and gastric carcinoma. METHODS: A rare case of malignant AN that initially manifested in the oral cavity of a 73-year-old patient is reported. RESULTS: A bladder and lung carcinoma were detected following the diagnosis of AN. CONCLUSIONS: The diagnostic importance of oral AN is emphasized because, in our patient, its recognition led to the detection of 2 occult malignant tumors.

TT virus infection of periodontal tissues: a controlled clinical and laboratory pilot study.

BACKGROUND: A novel single-strand, circular DNA virus has been recently isolated and named TT virus (TTV). It has been demonstrated that peripheral blood cells harbor TTV DNA, suggesting that the virus might replicate in lymphoid cells and contribute to lymphocyte imbalances with consequent immunosuppressive effects. The purpose of this study was to investigate the prevalence of TTV DNA in healthy and periodontally compromised subjects, evaluating the presence of the virus in the gingiva and saliva, and comparing virological results with clinical data. METHODS: Twenty-one patients (seven males and 14 females, aged 25 to 76 years) were enrolled in the study. Eleven subjects were diagnosed with moderate periodontitis, while 10 were periodontally healthy. A sample of saliva was taken from each patient before recording the periodontal data; subsequently, a gingival biopsy was performed. A real-time polymerase chain reaction was used to quantify the presence of TTV DNA in saliva and gingival specimens. RESULTS: A statistically significant association was found between TTV in gingival tissue and the presence of periodontitis (P = 0.0351), while no association was observed between TTV in saliva and the presence of periodontitis (P = 0.4762). CONCLUSIONS: A new DNA virus (TTV) was first identified in the gingival tissue and was found to be significantly associated with the presence of periodontitis. These findings need to be investigated in further studies.

Retrospective evaluation of the influence of the interleukin-1 genotype on radiographic bone levels in treated periodontal patients over 10 years.

BACKGROUND: A difference in genetic susceptibility to plaque accumulation has been advocated to explain different responses to periodontal therapy. The purpose of this study is to assess the role of the interleukin-1 (IL-1) polymorphism on the rate of bone and tooth loss in non-smoking periodontally treated patients during maintenance. METHODS: Sixty consecutive non-smoking patients (mean age 46.8 +/- 5.0) with moderate to severe periodontitis, treated and maintained for over 10 years were selected. At baseline (T0), radiographic evaluation (cemento-enamel junction [CEJ]-root apex, CEJ-bottom of defect mesial and distal, CEJ-bone crest mesial and distal, crown-root ratio) was performed. All patients received scaling and root planing; 36 patients then underwent surgical therapy. Subsequently, all patients were enrolled in a periodontal maintenance program with recall visits every 3.4 +/- 1.0 months for at least 10 years. At the latest recall visit (T2) the same radiographic measurements evaluated at baseline were taken and a DNA sample for IL-1 genetic susceptibility testing was collected and sent for analysis. RESULTS: Twenty-three of the 60 patients (38.3%) were IL-1 genotype positive. A total of 52 teeth (3.3%) out of 1,566 were lost due to periodontitis between T0 and T2; 28 of 957 (2.9%) in the IL-1 genotype negative group and 24 of 609 (3.9%) in IL-1 genotype positive group. The mean variation in bone defect level (DeltaBD) averaged -0.04 mm in IL-1 genotype negative patients and 0.01 mm in IL-1 genotype positive patients. The mean variation in bone crest level (DeltaBC) averaged -0.24 mm in IL-1 genotype negative patients and -0.28 mm in IL-1 genotype positive patients. However, a few patients showed significant differences in response to therapy based on initial bone levels and genotype. IL-1 negative patients who showed minimal initial bone loss responded to the therapy better than the IL-1 positive patients. IL-1 positive patients with severe initial bone loss showed a better response to the therapy than IL-1 negative patients. CONCLUSIONS: On average, there were no significant differences related to IL-1 genotype in tooth loss after 10 years in a non-smoking, well-maintained periodontal population. On an individual patient basis, the IL-1 genotype, in combination with the initial bone level, seems useful at the beginning of therapy for predicting bone level variation.

Coronally advanced flap procedure for root coverage. Treatment of root surface: root planning versus polishing.

This clinical study was designed to determine if mechanical instrumentation (root planing) of the exposed root is useful in treating gingival recession caused by traumatic toothbrushing following a coronally advanced flap (CAF). Ten patients with high levels of oral hygiene (full-mouth plaque score <20%), from 25 to 57 years of age, were selected for the study. Each patient showed 2 bilateral Class I or II maxillary recessions. A total of 20 recessions were treated. The difference in the recessions was < or =1 mm. In each patient, one recession was randomly assigned to the test group and the contralateral one to the control group. In the test group, the exposed root surface was polished at slow speed with a rubber cup and prophylaxis paste for 60 seconds. In the control group, the exposed root surface was planed with a sharp curet. In both test and control groups, a trapezoidal full- and partial-thickness flap was elevated, coronally displaced, and sutured to cover the treated root surface. Before treatment, the mean recession depth in the test group (polishing) was 3.1+/-1.1 mm; and in the control group (root planing), 2.9+/-1.0 mm. Three months after the described procedures, the test group (polishing) showed a mean recession reduction of 2.6+/-0.6 mm; mean percent root coverage was 89+/-14%. In the control group (root planing), the mean recession reduction was 2.3+/-0.7 mm and mean percent root coverage was 83+/-16%. The difference of recession reduction between the test and control group was not statistically significant (P = 0.1405), even though the test group showed slightly better clinical results in terms of root coverage. This prospective clinical, controlled, randomized study shows that mechanical instrumentation (root planing) of the exposed root surfaces is not necessary when shallow recessions caused by traumatic toothbrushing are treated using a coronally advanced flap (CAF) in patients with high levels of oral hygiene.