Accelerated Fatigue Model for Predicting Composite Restoration Failure.

An empirical method is proposed to predict the clinical performance of resin composite dental restorations by using laboratory data derived from simple specimens subjected to chemical degradation and accelerated cyclic fatigue. Three resin composites were used to fill dentin disks (2-mm inner diameter, 5-mm outer diameter, and 2 mm thick) made from bovine incisor roots. The specimens (n = 30 per group) were aged with different durations of a low-pH challenge (0, 24, and 48 h under pH 4.5) before being subjected to diametral compression with either a monotonically increasing load (fast fracture) or a cyclic load with a continuously increasing amplitude (accelerated fatigue). The data from 1 material were used to establish the relationship between laboratory time (number of cycles) and clinical time to failure (years) via the respective survival probability curves. The temporal relationship was then used to predict the clinical rates of failure for restorations made of the other 2 materials, and the predictions were compared with the clinical data to assess their accuracy. Although there were significant differences in the fast fracture strength among the groups of materials or durations of chemical challenge, fatigue testing was much better at separating the groups. Linear relationships were found between the laboratory and clinical times to failure for the first material (R2 = 0.90, 0.90, and 0.62 for the 0-, 24-, and 48-h low-pH groups, respectively). The clinical life of restorations made of the other 2 materials was best predicted with data from the 48-h low-pH groups. In conclusion, an accelerated fatigue model was successfully calibrated and applied to predict the clinical failure of resin composite restorations, and the predictions based on data obtained from chemically aged specimens provided the best agreement with clinical data.

Validation of finite element models for orthodontic aligners.

PURPOSE: Clear thermoplastic aligners have become popular in orthodontics, but the biomechanics of these devices is not well understood. Neither is the tooth movement induced by such devices. The aim of this study was to develop and validate finite element (FE) models for clear thermoplastic teeth aligners for orthodontic force prediction. METHODS AND MATERIALS: FE models were created from Micro-CT scans of an aligner and a model arch of teeth with one of the incisors tipped buccal-lingually by 2.4°. The models were uniformly meshed with 0.3-mm long elements. Linear-elastic mechanical properties provided by the material manufacturers were used. Fitting of the two components was simulated using Abaqus's interference fit, followed by frictional surface-to-surface interaction. The assembled FE model was validated by comparing its prediction for the teeth-aligner gaps and aligner surface strains with experimental data. The experimental teeth-aligner gaps were obtained from the Micro-CT scans whereas the aligner surface strains were measured using a 2-camera digital image correlation (DIC) system. RESULTS: Good agreement between prediction and measurement was obtained for both the teeth-aligner gaps and aligner surface strains. The linear regression between prediction and measurement for teeth-aligner gaps sampled at different positions had a R2 value of 0.99. The mean difference between prediction and measurement for the aligner surface strains (von Mises) over 1544 nodes on the labial side and 1929 nodes on the lingual side was 0.07% and 0.01%, respectively, both being lower than the mean background noise. CONCLUSION: A FE model for clear thermoplastic teeth aligners has been successfully developed and validated. The model can therefore be used with confidence to predict the forces and moments applied to teeth by the aligners, thus improving our understanding of the biomechanics of such devices and the tooth movement they induce.

Whole-genome and transcriptome analysis enhances precision cancer treatment options.

BACKGROUND: Recent advances are enabling delivery of precision genomic medicine to cancer clinics. While the majority of approaches profile panels of selected genes or hotspot regions, comprehensive data provided by whole-genome and transcriptome sequencing and analysis (WGTA) present an opportunity to align a much larger proportion of patients to therapies. PATIENTS AND METHODS: Samples from 570 patients with advanced or metastatic cancer of diverse types enrolled in the Personalized OncoGenomics (POG) program underwent WGTA. DNA-based data, including mutations, copy number and mutation signatures, were combined with RNA-based data, including gene expression and fusions, to generate comprehensive WGTA profiles. A multidisciplinary molecular tumour board used WGTA profiles to identify and prioritize clinically actionable alterations and inform therapy. Patient responses to WGTA-informed therapies were collected. RESULTS: Clinically actionable targets were identified for 83% of patients, of which 37% of patients received WGTA-informed treatments. RNA expression data were particularly informative, contributing to 67% of WGTA-informed treatments; 25% of treatments were informed by RNA expression alone. Of a total 248 WGTA-informed treatments, 46% resulted in clinical benefit. RNA expression data were comparable to DNA-based mutation and copy number data in aligning to clinically beneficial treatments. Genome signatures also guided therapeutics including platinum, poly-ADP ribose polymerase inhibitors and immunotherapies. Patients accessed WGTA-informed treatments through clinical trials (19%), off-label use (35%) and as standard therapies (46%) including those which would not otherwise have been the next choice of therapy, demonstrating the utility of genomic information to direct use of chemotherapies as well as targeted therapies. CONCLUSIONS: Integrating RNA expression and genome data illuminated treatment options that resulted in 46% of treated patients experiencing positive clinical benefit, supporting the use of comprehensive WGTA profiling in clinical cancer care.

A Review of Mechano-Biochemical Models for Testing Composite Restorations.

Due to the severe mechano-biochemical conditions in the oral cavity, many dental restorations will degrade and eventually fail. For teeth restored with resin composite, the major modes of failure are secondary caries and fracture of the tooth or restoration. While clinical studies can answer some of the more practical questions, such as the rate of failure, fundamental understanding on the failure mechanism can be obtained from laboratory studies using simplified models more effectively. Reviewed in this article are the 4 main types of models used to study the degradation of resin-composite restorations, namely, animal, human in vivo or in situ, in vitro biofilm, and in vitro chemical models. The characteristics, advantages, and disadvantages of these models are discussed and compared. The tooth-restoration interface is widely considered the weakest link in a resin composite restoration. To account for the different types of degradation that can occur (i.e., demineralization, resin hydrolysis, and collagen degradation), enzymes such as esterase and collagenase found in the oral environment are used, in addition to acids, to form biochemical models to test resin-composite restorations in conjunction with mechanical loading. Furthermore, laboratory tests are usually performed in an accelerated manner to save time. It is argued that, for an accelerated multicomponent model to be representative and predictive in terms of both the mode and the speed of degradation, the individual components must be synchronized in their rates of action and be calibrated with clinical data. The process of calibrating the in vitro models against clinical data is briefly described. To achieve representative and predictive in vitro models, more comparative studies of in vivo and in vitro models are required to calibrate the laboratory studies.

Laboratory simulation of longitudinally cracked teeth using the step-stress cyclic loading method.

AIM: To simulate in a laboratory setting longitudinal cracking in root filled premolar teeth, using cyclic mechanical fatigue. METHODOLOGY: Mesial-occlusal-distal (MOD) cavities were prepared in twenty root filled, single-rooted, mandibular premolars restored with fibre posts and resin composites. The samples were randomly divided into two groups based on the loading approaches: static loading with a crosshead speed of 0.5 mm/min and step-stress cyclic loading (1 Hz) with increasing amplitude. The loads and numbers of cycles to failure were recorded. Micro-CT was also used to identify the fracture modes. Statistical analysis was performed using Student's t-test. The level of significance was set at 0.05. RESULTS: The mean fracture loads for the static loading and cyclic loading groups were 769 ± 171 N and 720 ± 92 N, respectively. There was no significant difference between the two groups (P > 0.05). The proportions of longitudinal, cuspal and mixed-mode fractures under cyclic loading were 50%, 20% and 30%, respectively. Longitudinal fractures occurred with larger numbers of cycles and higher average loads per cycle compared with the other fractures. Static loading produced only cuspal fractures. CONCLUSIONS: Longitudinally cracked premolar teeth with root fillings were successfully produced using the step-stress cyclic loading method. This provides a more clinically representative methodology for studying cracked teeth in a laboratory setting.

Focal choroidal excavation in patients with central serous chorioretinopathy.

PURPOSE: To evaluate the prevalence and clinical features of focal choroidal excavation (FCE) in patients presenting with central serous chorioretinopathy (CSC). METHODS: This is a retrospective consecutive case series of consecutive patients with CSC who were referred for spectral domain optical coherence tomography (SD-OCT) between January 2010 and December 2011. Medical records were reviewed and clinical features including presence of FCE in SD-OCT, fluorescence angiography (FA), and indocyanine green angiography (ICGA) were studied. RESULTS: Among the 116 CSC patients assessed, FCE was found in 11 eyes of 7 (6.0%) patients. FCE was associated with subretinal fluid in six eyes of six patients and serous pigment epithelial detachment in three eyes of two patients. The mean central subfield retinal thickness of CSC eyes with FCE was 283.7 µm, compared with 377.5 µm for CSC eyes without FCE (Mann-Whitney U-test, P=0.020). Five FCE eyes of five patients had focal leakage on FA. Choroidal hyperpermeability on ICGA was found in seven CSC eyes with FCE, with four eyes showing hypofluorescent spot corresponding to the FCE. After a mean follow-up of 16 months, visual acuity of all 11 eyes with FCE remained stable or improved at the last follow-up. CONCLUSION: FCE is not an uncommon feature in patients with CSC and might be associated with choroidal hemodynamic disturbances.

Degradation in the dentin-composite interface subjected to multi-species biofilm challenges.

Oral biofilms can degrade the components in dental resin-based composite restorations, thus compromising marginal integrity and leading to secondary caries. This study investigates the mechanical integrity of the dentin-composite interface challenged with multi-species oral biofilms. While most studies used single-species biofilms, the present study used a more realistic, diverse biofilm model produced directly from plaques collected from donors with a history of early childhood caries. Dentin-composite disks were made using bovine incisor roots filled with Z100(TM) or Filtek(TM) LS (3M ESPE). The disks were incubated for 72 h in paired CDC biofilm reactors, using a previously published protocol. One reactor was pulsed with sucrose, and the other was not. A sterile saliva-only control group was run with sucrose pulsing. The disks were fractured under diametral compression to evaluate their interfacial bond strength. The surface deformation of the disks was mapped using digital image correlation to ascertain the fracture origin. Fracture surfaces were examined using scanning electron microscopy/energy-dispersive X-ray spectroscopy to assess demineralization and interfacial degradation. Dentin demineralization was greater under sucrose-pulsed biofilms, as the pH dropped <5.5 during pulsing, with LS and Z100 specimens suffering similar degrees of surface mineral loss. Biofilm growth with sucrose pulsing also caused preferential degradation of the composite-dentin interface, depending on the composite/adhesive system used. Specifically, Z100 specimens showed greater bond strength reduction and more frequent cohesive failure in the adhesive layer. This was attributed to the inferior dentin coverage by Z100 adhesive, which possibly led to a higher level of chemical and enzymatic degradation. The results suggested that factors other than dentin demineralization were also responsible for interfacial degradation. A clinically relevant in vitro biofilm model was therefore developed, which would effectively allow assessment of the degradation of the dentin-composite interface subjected to multi-species biofilm challenge.

A reproducible oral microcosm biofilm model for testing dental materials.

AIMS: Most studies of biofilm effects on dental materials use single-species biofilms, or consortia. Microcosm biofilms grown directly from saliva or plaque are much more diverse, but difficult to characterize. We used the Human Oral Microbial Identification Microarray (HOMIM) to validate a reproducible oral microcosm model. METHODS AND RESULTS: Saliva and dental plaque were collected from adults and children. Hydroxyapatite and dental composite discs were inoculated with either saliva or plaque, and microcosm biofilms were grown in a CDC biofilm reactor. In later experiments, the reactor was pulsed with sucrose. DNA from inoculums and microcosms was analysed by HOMIM for 272 species. Microcosms included about 60% of species from the original inoculum. Biofilms grown on hydroxyapatite and composites were extremely similar. Sucrose pulsing decreased diversity and pH, but increased the abundance of Streptococcus and Veillonella. Biofilms from the same donor, grown at different times, clustered together. CONCLUSIONS: This model produced reproducible microcosm biofilms that were representative of the oral microbiota. Sucrose induced changes associated with dental caries. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first use of HOMIM to validate an oral microcosm model that can be used to study the effects of complex biofilms on dental materials.

Quantifying dental biofilm growth using cross-polarization optical coherence tomography.

AIMS: Quantifying the ex vivo growth of complex multispecies dental biofilms using cross-polarization 1310-nm optical coherence tomography (CP-OCT) system was investigated. METHODS AND RESULTS: Bacterial microcosms, which were derived from plaque samples of paediatric subjects, were incubated in a biofilm reactor system containing discs of different dental materials for 72 h with daily sucrose pulsing (5×). CP-OCT analysis of biofilm mass was validated with crystal violet (CV) assays at various growth stages of these complex biofilms. CP-OCT was able to filter out the back-reflected signals of water layers in the hydrated biofilm and allowed for direct biofilm quantification. The overall depth-resolved scattering intensity of the biofilm showed very strong positive correlation with CV assay quantification (Spearman's ρ = 0.92) during the growth phase of the biofilm. CONCLUSION: CP-OCT was able to quantify the mass of the biofilm by measuring the overall depth-resolved scattering of the biofilm. SIGNIFICANCE AND IMPACT OF THE STUDY: CP-OCT has the ability to nondestructively monitor biofilm growth and elucidate the growth characteristics of these microcosms on different dental material compositions.

19p13.2 microduplication causes a Sotos syndrome-like phenotype and alters gene expression.

Up to 90% of individuals affected by Sotos syndrome have a pathogenic alteration of NSD1 (encodes nuclear receptor-binding Su-var, enhancer of zeste, and trithorax domain protein 1), a histone methyltransferase that functions as both a transcriptional activator and a repressor. Genomic copy number variations may also cause a Sotos-like phenotype. We evaluated a three-generation family segregating a Sotos-like disorder characterized by typical facial features, overgrowth, learning disabilities, and advanced bone age. Affected individuals did not have a detectable NSD1 mutation, but rather were found to have a 1.9 Mb microduplication of 19p13.2 with breakpoints in two highly homologous Alu elements. Because the duplication included the DNA methyltransferase gene (DNMT1), we assessed DNA methylation of peripheral blood and buccal cell DNA and detected no alterations. We also examined peripheral blood gene expression and found evidence for increased expression of genes within the duplicated region. We conclude that microduplication of 19p13.2 is a novel genomic disorder characterized by variable neurocognitive disability, overgrowth, and facial dysmorphism similar to Sotos syndrome. Failed compensation of gene duplication at the transcriptional level, as seen in peripheral blood, supports gene dosage as the cause of this disorder.

Osteomalacia: a case series of patients with atypical clinical orthopaedic presentations.

Osteomalacia is uncommon in an affluent subtropical city like Hong Kong, where sunlight exposure is adequate and nutritional support is good. We present three patients who had osteomalacia with different presentations. A 74-year-old male with oncogenic osteomalacia presented with multiple bone pain. His biochemical markers returned to normal 4 days postoperatively after resection of a second toe giant cell tumour of tendon sheath. A 62-year-old woman with a history of liver problem and proximal muscle weakness was admitted with atraumatic fracture of the left distal humerus due to osteomalacia. An 81-year-old vegetarian woman with inadequate sun exposure complained of multiple bone pains. Subsequent investigation revealed dietary- and sunlight-deficient osteomalacia with multiple bony abnormalities including marked femur bowing.

Strengthening of a model composite restoration using shape optimization: a numerical and experimental study.

OBJECTIVE: This study aims to validate a cavity shape optimization approach for improving the debonding resistance of dental restorations by carrying out fracture tests on restored model teeth with standard and optimized cavity designs. METHOD: The bio-mimetic stress-induced material transformation (SMT) optimization method was incorporated into the finite element (FE) program ABAQUS as a user material (UMAT) subroutine. The method uses stress minimization to optimize the cavity shape of a MOD restoration in an artificial premolar with special reference to the tooth-restoration interface under occlusal loads. The mechanical performance of the optimized design was first verified through FE analysis and then compared with that of the conventional design using fracture tests on model teeth. RESULTS: The SMT optimization process indicated a T-shape cavity as a more favorable design for the MOD restoration in the artificial premolar. Compared with the conventional parallel wall, or undercut design, the T-shape cavity was shown numerically to reduce the interfacial stresses by up to 69%, and experimentally to increase the mean debonding resistance of the model teeth by 23% (p<0.05). SIGNIFICANCE: Cavity shape optimization can help increase the debonding resistance of restored teeth by reducing the interfacial stresses between tooth and restoration under occlusal loads.

Risk factors for clustering of tuberculosis cases: a systematic review of population-based molecular epidemiology studies.

BACKGROUND: Many molecular epidemiology studies have been conducted to identify risk factors for clustering of tuberculosis (TB) cases in the population. OBJECTIVE: To estimate the impact of commonly investigated risk factors on TB clustering. METHODS: Ten electronic databases were searched up to January 2006 along with a hand search of the International Journal of Tuberculosis and Lung Disease and bibliographies of review articles. Meta-analyses of odds ratios (ORs) for various risk factors were conducted using random effect models, stratified by TB incidence. Meta-regressions were employed to account for the heterogeneity in clustering proportions and the magnitudes of risk. FINDINGS: The TB clustering proportion varied greatly (7.0-72.3%) among 36 studies in 17 countries. In multiple meta-regression analyses, high TB incidence, mean cluster size and conventional contact tracing were significantly associated with higher clustering. The pooled ORs (95%CIs) for low and high/intermediate TB incidence studies, using a cut off of 25/100000 per year, were 3.4 (2.7- 4.2) and 1.6 (1.3-2.1) for local-born status, 1.6 (1.5-1.7) and 1.7 (1.3-2.2) for pulmonary TB and 1.2 (1.1-1.3) and 1.3 (1.1-1.7) for smear-positive cases, respectively. Male sex, local birth, alcohol abuse and injection drug use were significantly higher risks in low TB incidence studies than in the high/intermediate ones. INTERPRETATION: Meta-analyses yielded significant estimates of ORs for several risk factors across both levels of TB incidence. Alcohol abuse, injection drug use and homelessness--all characteristics of marginalized populations--were found to be consistently significant in populations of low TB incidence. More research is needed to better understand TB transmission dynamics in high-burden countries.

In the absence of dietary surveillance, chitosan does not reduce plasma lipids or obesity in hypercholesterolaemic obese Asian subjects.

OBJECTIVE: To investigate the effects of Absorbitol on body weight, anthropometry, body composition, blood pressures and lipid profiles in obese, hypercholesterolaemic subjects without dietary restriction. DESIGN: A randomised, double blind. Placebo-controlled study. SUBJECTS: Normal volunteers with no history of chronic illnesses (n=88) who were obese (body fat percentage > 20% in males and > 30% in females) and hypercholesterolaemic (total cholesterol > 5.20 mmol/L). Sixty-eight (72.3%) subjects completed the study. INTERVENTION: After a 4 week run in phase, 4 placebo/Absorbitol (250 mg) capsules were prescribed 3 times a day before meals. Subjects received written information on healthy lifestyle but there was no dietary restriction or monitoring. MAIN OUTCOME MEASURES: Weight, body mass index, lean body mass, waist, hip, blood pressure, fasting lipids and insulin levels were taken at baseline, 4th and 16th week of the study. STATISTICAL ANALYSIS PERFORMED: Analyses were on an intention-to-treat basis. Comparisons between groups were made using Student's t and Mann-Whitney tests for parametric and non-parametric data respectively. RESULTS: There was no significant change in the measured parameters in Absorbitol treated subjects compared to those on placebo, with exception of HDL-cholesterol which increased in the absorbitol group and decreased in the placebo group (p=0.048). The side effects of Absorbitol were also comparable to that of placebo. CONCLUSIONS: In the absence of dietary surveillance, Absorbitol does not bring about improvement in weight, anthropometry, body composition, blood pressure or lipid profile.

Phagosome formation in Paramecium: roles of somatic and oral cilia and of solid particles as revealed by video microscopy.

The roles of somatic and oral cilia and solid particles during digestive vacuole (DV) formation in Paramecium multimicronucleatum were investigated using video-enhanced and immunofluorescence microscopy. Membrane incorporation into DVs was found to increase linearly with increasing particle concentration. The rate of discoidal vesicle transport to the cytopharynx was not affected by particles, showing that particles are not required for membrane trafficking to the cytopharynx. However, the presence of particles leads to an increased membrane fusion between the cytopharyngeal membrane and the discoidal vesicles. When live cells lost their somatic cilia on the left-ventral side anterior to the oral region due to deciliation, membrane incorporation into newly formed DVs was strongly inhibited. Using video-enhanced microscopy, latex beads were seen to be loaded along the quadrulus on the dorsal surface of the buccal cavity, but few beads were seen next to the dorsal and ventral peniculi. Particle sequestration into a pre-formed nascent digestive vacuole (NDV) was studied in Triton X-100-permeabilized cells whose ciliary beating was reactivated by the addition of Mg-ATP. Both beat frequency and the percentage of cells containing bead-labeled NDV were dependent on the Mg-ATP concentration: the higher the beat frequency, the higher the percentage of cells with a bead-labeled NDV. These results suggest that ciliary beating is probably the only mechanism required for particle accumulation in the NDV, while a coordinated beating of the somatic cilia on the left-ventral side anterior to the oral region as well as the quadrulus moves particles into the NDV. The beating of the peniculi may somehow prevent the backward flow of particles out of the NDV.

A man with osteoblastic metastasis and hypocalcaemia.

We report a case of an 80-year-old man with osteoblastic metastases from advanced carcinoma of the prostate presenting with a grand mal seizure resulting from severe hypocalcaemia. He had low serum phosphate and ionised calcium levels, elevated serum skeletal alkaline phosphatase and intact parathormone levels. 99mTc radioisotope bone scan revealed a "super bone scan" suggestive of osteomalacia. The serum 1, 25-dihydroxycholecalciferol level was unexpectedly elevated. The biochemical abnormalities persisted despite high dose calcium replacement, but improved with supraphysiological doses of 1,25 (OH)2 vitamin D3 (Rocaltrol) therapy. We hypothesise that the hypocalcaemia in this patient was due to vitamin D resistance secondary to a humoral factor secreted by the tumour.

Body fat distribution and cardiovascular risk in normal weight women. Associations with insulin resistance, lipids and plasma leptin.

OBJECTIVE: To systematically examine the correlations between insulin resistance, plasma leptin concentration, obesity and the distribution of fat assessed by anthropometry and magnetic resonance imaging in Asian women. DESIGN: A cross sectional study of non-diabetic, normal weight women. SUBJECTS: Twenty-one healthy women aged 38.8 y (s.d. 11.7) and BMI 22.6 kg/m2 (s.d. 2.3). MEASUREMENTS: Intraperitoneal, retroperitoneal and subcutaneous abdominal fat volume was assessed by magnetic resonance imaging. Anthropometric data were collected. Total fat mass was assessed by bioelectric impedance analysis. Fasting serum lipids, insulin and plasma leptin were assayed. RESULTS: Generalized obesity correlated with subcutaneous abdominal fat mass (r=0.83, P<0.001), but not with intra-abdominal fat mass. Both intraperitoneal fat mass and retroperitoneal fat mass increased with age (r=0.58, P=0.005 and r=0. 612, P=0.003, respectively). Abdominal subcutaneous fat mass was the most important determinant of insulin resistance and plasma leptin. Of the serum lipids, only fasting triglyceride correlated significantly with the waist-to-hip ratio. CONCLUSIONS: It is possible that the large size of the subcutaneous depot compared to the intra-abdominal depot overwhelms any metabolic differences between adipose tissue from these two sites, resulting in the stronger correlation between insulin resistance and subcutaneous abdominal fat mass rather than intra-abdominal fat mass. On the other hand, the distribution of fat between subcutaneous fat depots may be important in the metabolic syndrome given the correlation of fasting triglyceride with waist to hip ratio but not with abdominal fat. However, the study population was small, younger and leaner compared to previous studies and we may not be able to generalize these results to all segments of the population. We confirm that subcutaneous fat mass is the major determinant of plasma leptin.

Membrane trafficking and processing in Paramecium.

Cellular membranes are made in a cell's biosynthetic pathway and are composed of similar biochemical constituents. Nevertheless, they become differentiated as membrane components are sorted into different membrane-limited compartments. We summarize the morphological and immunological similarities and differences seen in the membranes of the various interacting compartments in the single-celled organism, Paramecium. Besides the biosynthetic pathway, membranes of the regulated secretory pathway, endocytic pathway, and phagocytic pathway are highlighted. Paramecium is a multipolarized cell in the sense that several different pools of membrane-limited compartments are targeted for exocytosis at very specific sites at the cell surface. Thus, the method used by this cell to sort and package its membrane subunits into different compartments, the processes used to transport these compartments to specific locations at the plasma membrane and to other intracellular fusion sites, the processes of membrane retrieval, and the processes of membrane docking and fusion are reviewed. Paramecium has provided an excellent model for studying the complexities of membrane trafficking in one cell using both morphological and immunocytochemical techniques. This cell also promises to be a useful model for studying aspects of the molecular biology of membrane sorting, retrieval, transport, and fusion.