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.

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.

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.

[Periodontic-endodontic lesions: diagnostic and therapeutic indications]. / Lesioni endo parodontali. Indicazioni diagnostiche e terapeutiche.

The presence of connections between periodontium and endodontium can lead to the diffusion of an infection from one apparatus to another. The involvement of both periodontium and endodontium is defined as Combined Periodontic-Endodontic lesions. This definition is not based on the initial etiology of the lesion and either the endodontic or periodontal lesion may be the cause or the result of the other or both may develop independently. The lesions must be correctly diagnosed for the best therapeutic approach. The diagnosis is based on clinical symptoms and radiographic analysis; clinical signs must show the presence of periodontal probing and pulpal necrosis. Radiographic examination can confirm the involvement of both periodontium and endodontium only if the lesion is present on the mesial and distal part of the diseased tooth; in the case of a palatal/lingual or vestibular lesion such evidence will not be detectable. The therapeutic approach is always based on an initial endodontic treatment followed, if needed, by the proper periodontal treatment.

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.

Limits to the development of fast neuromuscular transmission in zebrafish.

Zebrafish embryos have small and slow miniature end-plate currents (mEPCs), whereas only a few days later larval mEPCs are an order of magnitude larger and faster, being among the fastest of all neuromuscular synapses. To identify the bases for these changes we compared, in embryos and larvae, the properties and distributions of acetylcholine (ACh) receptors (AChRs) and acetylcholinesterase (AChE) as well as the ultrastructure of the developing neuromuscular junctions (NMJs). To mimic synaptic release, patches of muscle membrane were exposed briefly (for 1 ms) to a saturating concentration (10 mM) of ACh. The AChR deactivation kinetics were twice as slow in embryos compared with larvae. In both embryos and larvae, AChRs demonstrated open channel block by millimolar ACh, and this was detected during mEPCs, indicating that a high concentration of ACh is released at immature and mature NMJs. AChR and AChE distributions were compared using the selective fluorescently conjugated labels alpha-bungarotoxin and fasciculin 2, respectively. In larvae, punctate AChR clusters were detected whereas junctional AChE staining was less intense than that found at adult NMJs. Transmission electron microscopy revealed immature nerve endings in embryos that were closely juxtaposed to the surrounding muscle cells, whereas mature larval NMJs had a wider synaptic cleft with a conspicuous basal lamina over a limited region of synaptic contact. Our results indicate that ACh is released at high concentrations at immature NMJs, but its clearance is prolonged and the AChRs are dispersed, resulting in a slow mEPC time course until a mature cleft appears with densely packed faster AChRs and abundant AChE.

[Diabetes mellitus as a risk factor for periodontitis]. / Il diabete mellito come fattore di rischio per la parodontite.

Diabetes mellitus is an important disease of the endocrine system. Many studies have associated this disease to the pathogenesis and the severity of periodontal disease. The aim of this article is to illustrate the relation between diabetes mellitus and periodontal disease. Many studies show an important association between diabetes and the pathogenesis of periodontal disease. Vascular changes caused by hyperglycemia are associated to the development of periodontal pathogens species. Moreover diabetics show an exacerbate host response with hyperproduction of inflammatory mediators and polymorphonuclear dysfunction. Diabetics with good metabolic control and patients with good oral hygiene show a reduced risk of periodontitis. In conclusion, diabetes mellitus (IDDM and NIDDM) is an important risk factor for periodontitis. Odds Ratio is 3. Diabetes mellitus determines changes in bacterial population and production of inflammatory mediators, and reduces the efficacy of the host response. Good controlled diabetes do not cause a major risk of periodontitis and improve the results of the periodontal therapy. Moreover periodontal therapy may reduce the request of insulin in diabetics. It is reasonable a two-ways relation between diabetes and periodontal disease.

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.

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.

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.

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.

Transglutaminase-mediated fibronectin multimerization in lung endothelial matrix in response to TNF-alpha.

Exposure of lung endothelial monolayers to tumor necrosis factor (TNF)-alpha causes a rearrangement of the fibrillar fibronectin (FN) extracellular matrix and an increase in protein permeability. Using calf pulmonary artery endothelial cell layers, we determined whether these changes were mediated by FN multimerization due to enhanced transglutaminase activity after TNF-alpha (200 U/ml) for 18 h. Western blot analysis indicated that TNF-alpha decreased the amount of monomeric FN detected under reducing conditions. Analysis of (125)I-FN incorporation into the extracellular matrix confirmed a twofold increase in high molecular mass (HMW) FN multimers stable under reducing conditions (P < 0.05). Enhanced formation of such HMW FN multimers was associated with increased cell surface transglutaminase activity (P < 0.05). Calf pulmonary artery endothelial cells pretreated with TNF-alpha also formed nonreducible HMW multimers of FN when layered on surfaces precoated with FN. Inhibitors of transglutaminase blocked the TNF-alpha-induced formation of nonreducible HMW multimers of FN but did not prevent either disruption of the FN matrix or the increase in monolayer permeability. Thus increased cell surface transglutaminase after TNF-alpha exposure initiates the enhanced formation of nonreducible HMW FN multimers but did not cause either the disruption of the FN matrix or the increase in endothelial monolayer permeability.

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.

Tissue engineering technology for gingival augmentation procedures: a case report.

Tissue engineering technology has been used in periodontal surgery. A patient who needed gingival augmentation prior to a single prosthetic restoration was treated by means of a tissue engineering technique. Results are presented in this case report.

Hepatic fibronectin matrix turnover in rats: involvement of the asialoglycoprotein receptor.

Fibronectin (Fn) is a major adhesive protein found in the hepatic extracellular matrix (ECM). In adult rats, the in vivo turnover of plasma Fn (pFn) incorporated into the liver ECM is relatively rapid, i.e., <24 h, but the regulation of its turnover has not been defined. We previously reported that cellular Fn (cFn) and enzymatically desialylated plasma Fn (aFn), both of which have a high density of exposed terminal galactose residues, rapidly interact with hepatic asialoglycoprotein receptors (ASGP-R) in association with their plasma clearance after intravenous infusion. With the use of adult male rats (250-350 g) and measurement of the deoxycholate (DOC)-insoluble (125)I-labeled Fn in the liver, we determined whether the ASGP-R system can also influence the hepatic matrix retention of various forms of Fn. There was a rapid deposition of (125)I-pFn, (125)I-aFn, and (125)I-cFn into the liver ECM after their intravenous injection. Although (125)I-pFn was slowly lost from the liver matrix over 24 h, more than 90% of the incorporated (125)I-aFn and (125)I-cFn was cleared within 4 h (P < 0.01). Intravenous infusion of excess nonlabeled asialofetuin to competitively inhibit the hepatic ASGP-R delayed the rapid turnover of both aFn and cFn already incorporated within the ECM of the liver. ECM retention of both (125)I-aFn and (125)I-cFn was also less than (125)I-pFn (P < 0.01) as determined in vitro using liver slices preloaded in vivo with either tracer form of Fn. The hepatic ASGP-R appears to participate in the turnover of aFn and cFn within the liver ECM, whereas a non-ASGP-R-associated endocytic pathway apparently influences the removal of normal pFn incorporated within the hepatic ECM, unless it becomes locally desialylated.

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.

Lung matrix incorporation of plasma fibronectin reduces vascular permeability in postsurgical bacteremia.

Plasma fibronectin (pFN) can incorporate into the lung extracellular matrix (ECM) as well as enhance hepatic cell phagocytic removal of bloodborne microparticulate debris that can contribute to lung vascular injury. Treatment of human pFN (hFN) with N-ethylmaleimide (NEM) blocks its ECM incorporation but not its ability to augment phagocytosis. Using hFN purified from fresh human plasma cryoprecipitate, we compared the effect of NEM-treated hFN versus normal hFN on lung transvascular protein clearance (TVPC) in postoperative bacteremic sheep to determine whether the ability of hFN to attenuate the increase in lung endothelial permeability required its ECM incorporation. Sheep with lung lymph fistulas were infused with a sublethal dose of Pseudomonas aeruginosa (5 x 10(8)) 48 h after surgery. In the first study, sheep received either FN-rich human cryoprecipitate, FN-deficient cryoprecipitate, FN purified from cryoprecipitate (hFN), FN-deficient cryoprecipitate reconstituted with purified hFN, or the sterile saline diluent. In the second study, sheep received either 200 mg of purified hFN (group I), 200 mg of NEM-treated hFN (group II), or the saline diluent (group III). In the first study, the increase in TVPC after bacterial challenge was attenuated by FN-rich cryoprecipitate, hFN, or reconstituted FN-deficient cryoprecipitate (P < 0.05) but not by saline and FN-deficient cryoprecipitate. In the second study, TVPC increased by 2 h (P < 0.05) and peaked over 4-8 h (P < 0.05) at 380-420% above baseline in postoperative bacteremic sheep given the diluent (group III). In contrast, intravenous infusion of hFN, but not of NEM-treated hFN, significantly (P < 0.05) attenuated this increase of lung protein clearance. Thus the ability for the intravenously infused purified pFN to attenuate the increase in lung endothelial protein permeability in sheep during postsurgical bacteremia appears to require its ECM incorporation into the interstitial ECM of the lung.

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.

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.

Circulating cellular fibronectin may be a natural ligand for the hepatic asialoglycoprotein receptor: possible pathway for fibronectin deposition and turnover in the rat liver.

It has been postulated that the in vivo removal of many plasma glycoproteins after desialylation is mediated by their interaction with a specific endocytic receptor on hepatocytes called the asialoglycoprotein receptor (ASGP-R), which is known to have a high affinity for specific carbohydrate residues, such as galactose. However, this mechanism has never been proven in vivo, nor has a naturally occurring ligand for the ASGP-R been identified. We investigated the influence of the terminal galactose residues on plasma fibronectin (pFn) on its liver deposition and turnover in adult rats, using neuraminidase to remove sialic acid residues to expose galactose residues. We also tested the hypothesis that the normal presence of a large amount of terminal galactose residues in cellular Fn (cFn) may allow cFn to serve as a natural ligand readily able to interact with the ASGP-R. In contrast to the slow clearance of normal pFn from the blood, cFn and desialylated pFn (aFn) displayed a rapid plasma clearance (P < .001) with greater than 50% of both the 125I-cFn or 125I-aFn depositing in the liver within 15 minutes. The enhanced plasma removal and liver deposition of both 125I-cFn and 125I-aFn was competitively inhibited (P < .01) by prior intravenous infusion of excess asialofetuin, which can selectively bind to the ASGP-R. The enzymatic addition of terminal sialic acid residues onto cFn to "mask" or "cap" the normally exposed galactose residues delayed the rapid plasma removal of cFn. Accelerated degradation of 125I-aFn and 125I-cFn as compared with 125I-pFn was demonstrated in vitro by both primary cultures of normal rat hepatocytes or incubated (37 degrees C) tissue slices of livers harvested from normal rats after in vivo preloading with tracer 125I-Fn forms. Thus, the ASGP-R appears to directly participate in the rapid in vivo removal of cFn from the blood, while native pFn may be removed by an alternative pathway unless it can become desialylated in vivo. These findings suggest that cFn may be a naturally occurring ligand that does not require desialylation before removal by the ASGP-R on hepatocytes.