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Studies of ripening cherry fruit suggest that pectin depolymerization does not affect fruit softening. The water- and acid-soluble fractions possess more neutral sugar besides than the oxalate-soluble fraction. The major changes are galatan degradation and absence of arabinan solubilization in acid-soluble pectic substance.

Statement of Problem: Analysis of the cost and time efficiency of removable complete dentures (RCDs) fabricated using conventional and computer-aided design and computer-aided manufacturing (CAD-CAM) processes remains limited., Purpose: The purpose of this systematic review was to evaluate the time efficiency and cost of conventional, hybrid, and completely digital workflows throughout the entire process of RCD fabrication., Material and Methods: Article selection followed the Preferred Reporting Items for Systematic Review and Meta-Analysis Protocols (PRISMA-P) 2015 guidelines and was registered with the international prospective register of systematic reviews (PROSPERO). The bibliographic search covered the PubMed and Cochrane databases using keywords related to removable complete dentures, digital workflows, and economic factors and covered articles from January 2010 to June 2024. In total, 1188 articles were identified and assessed. A bottom-up search identified an additional 10 articles through cited works. Data on cost and time analyses were reported in a qualitative synthesis., Results: The qualitative synthesis included 8 articles published between 2015 and 2024 showing that digital and hybrid workflows generally reduced both clinical and laboratory time and overall costs compared with conventional workflows. The hybrid protocol required significantly less clinical time, with reductions ranging from approximately 1 to 4 hours. Hybrid and completely digital workflows reduced laboratory manufacturing time by approximately 6 to 7 hours compared with conventional workflows. Cost analyses revealed that digital and hybrid workflows were more profitable and cost effective than conventional workflows., Conclusions: The findings suggest that digital and hybrid workflows for RCD fabrication are more time efficient and cost effective than conventional methods., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The pre-integration steps of the HIV-1 viral cycle are some of the most valuable targets of recent therapeutic innovations. HIV-1 integrase (IN) displays multiple functions, thanks to its considerable conformational flexibility. Recently, such flexible proteins have been characterized by their ability to form biomolecular condensates as a result of Liquid-Liquid-Phase-Separation (LLPS), allowing them to evolve in a restricted microenvironment within cells called membrane-less organelles (MLO). The LLPS context constitutes a more physiological approach to study the integration of molecular mechanisms performed by intasomes (complexes containing viral DNA, IN, and its cellular cofactor LEDGF/p75). We investigated here if such complexes can form LLPS in vitro and if IN enzymatic activities were affected by this LLPS environment. We observed that the LLPS formed by IN-LEDGF/p75 functional complexes modulate the in vitro IN activities. While the 3'-processing of viral DNA ends was drastically reduced inside LLPS, viral DNA strand transfer was strongly enhanced. These two catalytic IN activities appear thus tightly regulated by the environment encountered by intasomes., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Statement of Problem: Technological advances in digital acquisition tools have increased the scope of intraoral scanners (IOSs), including scanning a removable complete denture (RCD) to replicate it. However, studies assessing the accuracy of IOSs for replicating a maxillary or mandibular RCD are lacking., Purpose: The purpose of this in vitro study was to evaluate the accuracy (trueness and precision) of 3 IOSs while replicating a maxillary and mandibular RCD., Material and Methods: One maxillary and 1 mandibular RCD were scanned with a desktop scanner (D2000) to obtain the reference model. Two operators scanned each RCD 5 times with 3 different IOSs (TRIOS 4, Primescan, and IS3800), following a predefined acquisition protocol. The 60 study models obtained were compared with the reference model using the Geomagic software program. For each comparison, the mean and standard deviation of discrepancy were calculated. Distances were measured on both the reference and the study model, and differences were calculated to assess whether sagittal or transverse deformations were present. The tolerance percentage of the volume of the digital model compared with the volume of the reference model was determined (difference tolerance was set at 0.1 mm). A univariate analysis of variance followed by a post hoc analysis using the Student-Newman-Keuls (α=.05) test was performed to determine the truest and the most precise IOS., Results: The TRIOS 4 and Primescan IOSs had comparable trueness, with mean dimensional variations of 47 ±27 µm and 57 ±8 µm respectively compared with the reference model. The IS3800 had a lower trueness (98 ±35 µm). Primescan was significantly more precise with a mean standard deviation of 64 ±15 µm (P<.05). The TRIOS 4 (141 ±48 µm) and IS3800 (129 ±24 µm) had comparable precision. Primescan showed the least sagittal and transverse deformation., Conclusions: This study determined that an RCD can be replicated using an IOS, although all IOSs did not have equal accuracy. An in vivo study needs to assess whether this procedure is clinically acceptable., (Copyright © 2024 Editorial Council for The Journal of Prosthetic Dentistry. Published by Elsevier Inc. All rights reserved.)

Completion of neuronal migration is critical for brain development. Kif21b is a plus-end-directed kinesin motor protein that promotes intracellular transport and controls microtubule dynamics in neurons. Here we report a physiological function of Kif21b during radial migration of projection neurons in the mouse developing cortex. In vivo analysis in mouse and live imaging on cultured slices demonstrate that Kif21b regulates the radial glia-guided locomotion of newborn neurons independently of its motility on microtubules. We show that Kif21b directly binds and regulates the actin cytoskeleton both in vitro and in vivo in migratory neurons. We establish that Kif21b-mediated regulation of actin cytoskeleton dynamics influences branching and nucleokinesis during neuronal locomotion. Altogether, our results reveal atypical roles of Kif21b on the actin cytoskeleton during migration of cortical projection neurons., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

HIV-1 integrase-LEDGF allosteric inhibitors (INLAIs) share the binding site on the viral protein with the host factor LEDGF/p75. These small molecules act as molecular glues promoting hyper-multimerization of HIV-1 IN protein to severely perturb maturation of viral particles. Herein, we describe a new series of INLAIs based on a benzene scaffold that display antiviral activity in the single digit nanomolar range. Akin to other compounds of this class, the INLAIs predominantly inhibit the late stages of HIV-1 replication. A series of high-resolution crystal structures revealed how these small molecules engage the catalytic core and the C-terminal domains of HIV-1 IN. No antagonism was observed between our lead INLAI compound BDM-2 and a panel of 16 clinical antiretrovirals. Moreover, we show that compounds retained high antiviral activity against HIV-1 variants resistant to IN strand transfer inhibitors and other classes of antiretroviral drugs. The virologic profile of BDM-2 and the recently completed single ascending dose phase I trial (ClinicalTrials.gov identifier: NCT03634085) warrant further clinical investigation for use in combination with other antiretroviral drugs. Moreover, our results suggest routes for further improvement of this emerging drug class.

RNA can regulate its own synthesis without auxiliary proteins. For example, U-rich RNA sequences signal RNA polymerase (RNAP) to pause transcription and are required for transcript release at intrinsic terminators in all kingdoms of life. In contrast, the regulatory RNA putL suppresses pausing and termination in cis. However, how nascent RNA modulates its own synthesis remains largely unknown. We present cryo-EM reconstructions of RNAP captured during transcription of putL variants or an unrelated sequence at a U-rich pause site. Our results suggest how putL suppresses pausing and promotes its synthesis. We demonstrate that transcribing a U-rich sequence, a ubiquitous trigger of intrinsic termination, promotes widening of the RNAP nucleic-acid-binding channel. Widening destabilizes RNAP interactions with DNA and RNA to facilitate transcript dissociation reminiscent of intrinsic transcription termination. Surprisingly, RNAP remains bound to DNA after transcript release. Our results provide the structural framework to understand RNA-mediated intrinsic transcription termination., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 Elsevier Inc. All rights reserved.)

This case report presents the treatment of a 33-year-old patient with a genetic, generalized microdontia of permanent teeth. Microdontia is a developmental dental anomaly of the teeth characterized by a decrease in their size. In the literature, treatment has been multidisciplinary, often utilizing orthodontics and implantology. However, for adult patients with generalized microdontia who do not benefit from these treatments as much, a combination of adhesive dentistry, endodontics and removable prostheses remains a therapeutic alternative to consider. Given the specificities of the patient and the nature of the care, the objective of this treatment option was to manage the patient's comfort while guaranteeing quality of care. A feature of this treatment was the use of general anesthesia for endodontic treatments and exodontia. Chairside CAD/CAM and adhesive dentistry reduced the chairside time and preserved healthy dental tissue.

Purpose: We examined the impact of matrix food structure on post-prandial folate bioavailability (and other macronutrients) in human volunteers using a randomized, controlled, crossover experimental design., Methods: Twelve healthy male volunteers (22.6 ± 0.4 years old) were offered four food models (differing in matrix structure: Custard, Pudding, Sponge cake and Biscuit) to which 1 mg of folic acid was added, according to a randomized, controlled, crossover experimental design. Plasma folates, glucose, insulin, alpha amino nitrogen and triglycerides were measured over the post-prandial period (from T0 to T480 min)., Results: Food matrix structure was capable of altering folate plasma availability. The highest folate availability was observed for pudding and to a lesser extent Sponge cake whereas the lowest was for the two matrices presenting extreme rheological properties: Custard (liquid) (P < 0.05 total AUC) and to a lesser extent Biscuit (hard solid) (P < 0.05, AUC 180 min). The analysis of plasma kinetics of appearance of other nutrients/metabolites helps to understand/explain the lower bioavailability of folates in Custard and Biscuit., Conclusion: A least overall efficient bio-accessibility of all macronutrients and folic acid is observed in the gut lumen for Biscuit (delayed/incomplete destructuration of biscuit along the digestive tract). On the contrary, the lower folic acid absorption observed with custard does not fit with the rapid plasma appearance of other nutrients and should require further investigation.

The synthesis of acyl fluorides using the deoxofluorination reaction of carboxylic acids using XtalFluor-E is described. This transformation, assisted by a catalytic amount of NaF, occurs at room temperature in EtOAc, where XtalFluor-E behaves as the activating agent and the fluoride source. A wide range of acyl fluorides were obtained in moderate to excellent yields (36-99%) after a simple filtration on a pad of silica gel. We also demonstrated that sequential deoxofluorination/amidation was possible.

The distribution of food particles in a chewed bolus characterizes the food destruction after food oral processing (FOP). Previous reviews report that it could be affected by a lot of parameters as the number of chewing strokes, the dental status, but the conditions for producing reproducible data allowing inter-studies comparison have not been clearly described yet. This systematic review aims to identify the variables that can affect bolus granulometry determination, and to calculate their relative weights in the median particle size (D 50 ) variations. The systematic review focuses on granulometry expressed as D50 of the most used foods (peanuts, carrots, and almonds) and materials (Optosil and Optocal). Based on 58 studies, 5 variables among 60 being extracted could explain the D 50 variations. Conceptual differences between the conditions for FOP should be considered. After Chewing-test, the bolus is collected after a predefined number of strokes and its granulometry characterizes the effects of the dental and muscular apparatus on food destruction, while after Mastication-test the bolus is collected at the swallowing threshold, and its granulometry reflects the outcome of the abilities of the subject to adapt his/her mastication behavior to food texture. Experimental conditions related either to physical sieving or image analysis used to analyze the collected boluses impact the D 50 values. Finally, when type of test, sieving conditions, type of food or material, number of chewing strokes, and the oral status of the subjects are controlled, mean D 50 values are reproducible and could be used for inter-studies comparisons. PRACTICAL APPLICATIONS: This review provides tables that could be useful to control mean D 50 variations in further research., (© 2018 Wiley Periodicals, Inc.)

A new methodology to access enantiopure α,α-difluoromethyl alcohols is hereby being described. The strategy relies on the use of an enantiopure aryl α,α-difluoromethyl sulfoxide employed as chiral and removable auxiliary for the stereoselective difluoromethylation of carbonyl derivatives. The obtained α,α-difluoro-β-hydroxysulfoxides displayed unprecedented diastereomeric ratios.

Type 2 DNA topoisomerases (Top2) are critical components of key protein complexes involved in DNA replication, chromosome condensation and segregation, as well as gene transcription. The Top2 were found to be the main targets of anticancer agents, leading to intensive efforts to understand their functional and physiological role as well as their molecular structure. Post-translational modifications have been reported to influence Top2 enzyme activities in particular those of the mammalian Top2α isoform. In this study, we identified phosphorylation, and for the first time, acetylation sites in the human Top2α isoform produced in eukaryotic expression systems. Structural analysis revealed that acetylation sites are clustered on the catalytic domains of the homodimer while phosphorylation sites are located in the C-terminal domain responsible for nuclear localization. Biochemical analysis of the eukaryotic-specific K168 residue in the ATPase domain shows that acetylation affects a key position regulating ATP hydrolysis through the modulation of dimerization. Our findings suggest that acetylation of specific sites involved in the allosteric regulation of human Top2 may provide a mechanism for modulation of its catalytic activity.

Background: Historically, the complete removable denture is the last prosthetic procedure to switch to digital techniques whose advantages are mainly observed in the laboratory stages; however, it is not possible to measure the depressibility of the oral mucosa using optical cameras, thus conventional impression techniques are still necessary. This article describes the clinical and laboratory procedure and practitioners appraisal of the first fifteen digitally designed complete removable dental prostheses., Methods: Several systems are now available including the Wieland® Digital Denture® which offers a complete procedure. This system is composed of a five axis-milling machine combined with a laboratory scanner and a design software application. Fifteen rehabilitations were carried out using the Wieland® system., Results: The practitioner's role is simplified by intraoral recording with a central point and a reduced number of sessions. The prosthesis laboratory requires considerable investment in learning and equipment, making it possible to obtain ideal mounting assemblies in accordance with the occluso-prosthetic concept of bilateral balanced occlusion. The absence of polymerization and therefore of base deformation risks reduce the equilibration step. Finally, the creation of templates as an alternative to the assembly of teeth on wax makes it possible to functionally validate (masticatory and phonatory) the future dentures. However, this procedure still presented some limitations in terms of scanning and software scope of applications., Conclusion: Digital denture design software is relatively efficient and helps to standardize clinical results. However, to this date, improvements of the software are still required for a routine use.

The frequent instability of mandibular removable complete dentures affects patient Oral Health Related Quality of Life (OHRQoL). An innovative therapeutic strategy used to improve stability involves placing four symphyseal mini-implants. This study was aimed at assessing OHRQoL over time in subjects in which mini-implants were placed and exploring if certain parameters could predict the evolution of their OHRQoL. The OHRQoL of subjects with dentures was assessed using the Geriatric Oral Health Assessment Index (GOHAI) before (T0), 2-6 months (T1), twelve months (T2) and twenty-four or more months (T3) after mini-implant setting. Age, gender and chewing ability were tested as explanatory variables for the change in OHRQoL with time. Thirteen women and six men were included (mean age: 69 ± 10 years). After treatment, mean GOHAI scores at T1, T2 and T3 increased significantly ( p < 0.001). The GOHAI-Add mean score was not affected by age or gender. Baseline chewing ability impacted the "functional" and "pain and discomfort" fields of the mean GOHAI scores ( p < 0.05). The OHRQoL quickly improved after mini-implant placement in complete denture wearers and then stabilized over time. Baseline chewing ability can be used as a predictive parameter of OHRQoL., Competing Interests: The authors declare no conflict of interest.

The fluoride-promoted vinylogous Mukaiyama-Michael reaction of 2-[(trimethylsilyl)oxy]furan with diverse α,β-unsaturated ketones is described. The TBAF-catalyzed VMMR afforded high anti-diastereoselectivity irrespective of the solvents used. The KF/crown ethers catalytic systems proved to be highly efficient in terms of yields and resulted in a highly diastereoselective unprecedented solvent/catalyst switchable reaction. Anti-adducts were obtained as single diastereomers or with excellent diastereoselectivities when benzo-15-crown-5 in CH 2 Cl 2 was employed. On the other hand, high syn-diastereoselectivities (from 73:27 to 96:4) were achieved by employing dicyclohexane-18-crown-6 in toluene. On the basis of DFT calculations, the catalysts/solvent-dependent switchable diastereoselectivities are proposed to be the result of loose or tight cation-dienolate ion pairs.

Background: The renewal of removable dentures is often suggested to denture wearers subject to discomfort. However, the impact of this rehabilitation on patients' oral health related quality of life and their removable dentures related satisfaction is still unknown. This study was aimed at assessing these patient-centered outcomes and the potential impact of different factors., Methods: A cohort of 116 patients in need of removable dental prostheses rehabilitation was recruited at a dental hospital over a period of 1 year. The subjects were separated into two groups according to their prosthesis experience (group in need of removable dentures renewal/group needing an removable dentures for the first time). Subjects were asked to answer the "Geriatric Oral Health Assessment Index" (GOHAI) and the "McGill Denture Satisfaction Instrument" before and after a prosthesis integration period (9-12 weeks)., Results: GOHAI scores were slightly higher for patients with removable dentures renewal (from 40.6 ± 10.3 to 47.1 ± 10.0, p < 0.001), independently of the type of prosthetic rehabilitation. However, the scores of the GOHAI functional field did not change. Subjects with no removable dentures experience presented an increase in their functional GOHAI score (p < 0.001). Regarding patient removable dentures related satisfaction, only the "Esthetic" (p < 0.001), "Chewing efficiency" (p < 0.001) and "Oral condition" (p < 0.01) items increased after prosthesis renewal., Conclusions: This study showed that renewing removable dentures only moderately improved the oral health related quality of life and removable dentures related satisfaction of patients, regardless of age, gender or type of rehabilitation. Other tasks are necessary such as the analysis of physiological parameters and qualitative research on patient's expectations.

Antibody-drug conjugates (ADC), combining the specificity of tumor recognition by monoclonal antibodies (mAb) and the powerful cytotoxicity of anticancer drugs, are currently under growing interest and development. Here, we studied the potential of Chi-Tn, a mAb directed to a glyco-peptidic tumor-associated antigen, to be used as an ADC for cancer treatment. First, we demonstrated that Chi-Tn specifically targeted tumor cells in vivo. Also, using flow cytometry and deconvolution microscopy, we showed that the Chi-Tn mAb is rapidly internalized - condition necessary to ensure the delivery of conjugated cytotoxic drugs in an active form, and targeted to early and recycling endosomes. When conjugated to saporin (SAP) or to auristatin F, the Chi-Tn ADC exhibited effective cytotoxicity to Tn-positive tumor cells in vitro, which correlated with the level of tumoral Tn expression. Furthermore, the Chi-Tn mAb conjugated to auristatin F also exhibited efficient antitumor activity in vivo, validating for the first time the use of an anti-Tn antibody as an effective ADC.

A key challenge in anticancer therapy is to gain control over the biodistribution of cytotoxic drugs. The most promising strategy consists in conjugating drugs to tumor-targeting carriers, thereby combining high cytotoxic activity and specific delivery. To target Gb3-positive cancer cells, we exploit the non-toxic B-subunit of Shiga toxin (STxB). Here, we have conjugated STxB to highly potent auristatin derivatives (MMA). A former linker was optimized to ensure proper drug-release upon reaching reducing environments in target cells, followed by a self-immolation step. Two conjugates were successfully obtained, and in vitro assays demonstrated the potential of this targeting system for the selective elimination of Gb3-positive tumors., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

Crowns produced using CAD-CAM technology must be evaluated so they can be integrated in the dentist's therapeutic arsenal. To this end, an evaluation tool for the fixed element units was developed and tested in a randomized cross-over pilot study. At a hospital centre, 10 single crowns from the same dental preparation and produced using CEREC technology were compared to Ceramo-Metal Crowns produced using a conventional procedure. Practitioners generally evaluated CEREC crowns more positively, corroborating the subjects' preference for these crowns. The CEREC technology used in a hospital setting meets the requirements of both patients and practitioners.

Type 2A DNA topoisomerases (Topo2A) remodel DNA topology during replication, transcription and chromosome segregation. These multisubunit enzymes catalyze the transport of a double-stranded DNA through a transient break formed in another duplex. The bacterial DNA gyrase, a target for broad-spectrum antibiotics, is the sole Topo2A enzyme able to introduce negative supercoils. We reveal here for the first time the architecture of the full-length Thermus thermophilus DNA gyrase alone and in a cleavage complex with a 155 bp DNA duplex in the presence of the antibiotic ciprofloxacin, using cryo-electron microscopy. The structural organization of the subunits of the full-length DNA gyrase points to a central role of the ATPase domain acting like a 'crossover trap' that may help to sequester the DNA positive crossover before strand passage. Our structural data unveil how DNA is asymmetrically wrapped around the gyrase-specific C-terminal β-pinwheel domains and guided to introduce negative supercoils through cooperativity between the ATPase and β-pinwheel domains. The overall conformation of the drug-induced DNA binding-cleavage complex also suggests that ciprofloxacin traps a DNA pre-transport conformation.

Ribonuclease P (RNase P) and RNase MRP are closely related ribonucleoprotein enzymes, which process RNA substrates including tRNA precursors for RNase P and 5.8 S rRNA precursors, as well as some mRNAs, for RNase MRP. The structures of RNase P and RNase MRP have not yet been solved, so it is unclear how the proteins contribute to the structure of the complexes and how substrate specificity is determined. Using electron microscopy and image processing we show that eukaryotic RNase P and RNase MRP have a modular architecture, where proteins stabilize the RNA fold and contribute to cavities, channels and chambers between the modules. Such features are located at strategic positions for substrate recognition by shape and coordination of the cleaved-off sequence. These are also the sites of greatest difference between RNase P and RNase MRP, highlighting the importance of the adaptation of this region to the different substrates.

The CCR4-NOT complex is a deadenylation complex, which plays a major role for mRNA stability. The complex is conserved from yeast to human and consists of nine proteins NOT1-NOT5, CCR4, CAF1, CAF40 and CAF130. We have successfully isolated the complex using a Protein A tag on NOT1, followed by cross-linking on a glycerol gradient. All components of the complex were identified by mass spectrometry. Electron microscopy of negatively stained particles followed by image reconstruction revealed an L-shaped complex with two arms of similar length. The arms form an accessible cavity, which we think could provide an extensive interface for RNA-deadenylation., (Copyright © 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Nascent mRNAs produced by transcription in the nucleus are subsequently processed and packaged into mRNA ribonucleoprotein particles (messenger ribonucleoproteins (mRNPs)) before export to the cytoplasm. Here, we have used the poly(A)-binding protein Nab2 to isolate mRNPs from yeast under conditions that preserve mRNA integrity. Upon Nab2-tandem affinity purification, several mRNA export factors were co-enriched (Yra1, Mex67, THO-TREX) that were present in mRNPs of different size and mRNA length. High-throughput sequencing of the co-precipitated RNAs indicated that Nab2 is associated with the bulk of yeast transcripts with no specificity for different mRNA classes. Electron microscopy revealed that many of the mRNPs have a characteristic elongated structure. Our data suggest that mRNPs, although associated with different mRNAs, have a unifying core structure.

The genome of Mycoplasma pneumoniae is among the smallest found in self-replicating organisms. To study the basic principles of bacterial proteome organization, we used tandem affinity purification-mass spectrometry (TAP-MS) in a proteome-wide screen. The analysis revealed 62 homomultimeric and 116 heteromultimeric soluble protein complexes, of which the majority are novel. About a third of the heteromultimeric complexes show higher levels of proteome organization, including assembly into larger, multiprotein complex entities, suggesting sequential steps in biological processes, and extensive sharing of components, implying protein multifunctionality. Incorporation of structural models for 484 proteins, single-particle electron microscopy, and cellular electron tomograms provided supporting structural details for this proteome organization. The data set provides a blueprint of the minimal cellular machinery required for life.

Vacuolar ATPases (V-ATPases) are ATP-dependent proton pumps that maintain the acidity of cellular compartments. They are composed of a membrane-integrated proton-translocating V(0) and an extrinsic cytoplasmic catalytic domain V(1), joined by several connecting subunits. To clarify the arrangement of these peripheral connections and their interrelation with other subunits of the holocomplex, we have determined the solution structures of isolated EG and EGC connecting subcomplexes by small angle X-ray scattering and the 3D map of the yeast V-ATPase by electron microscopy. In solution, EG forms a slightly kinked rod, which assembles with subunit C into an L-shaped structure. This model is supported by the microscopy data, which show three copies of EG with two of these linked by subunit C. However, the relative arrangement of the EG and C subunits in solution is more open than that in the holoenzyme, suggesting a conformational change of EGC during regulatory assembly and disassembly.

Background: Changes in cell surface glycosylation that accompany apoptosis are thought to be involved in the recognition and removal of apoptotic cells by phagocytes, but in most instances these changes are ill defined. To improve our understanding of this phenomenon, we designed a trivariate flow cytometry procedure that allows direct comparison of cell surface glycosylation in apoptotic and viable cells., Methods: The annexin V/propidium iodide assay has been adapted for cell surface glycosylation analysis by combining the use of these two reagents with biotinylated lectins, and this has been used to investigate camptothecin-induced apoptosis in U-937 cells., Results: Although numerous lectins are potent inducers of apoptosis, we found that it is possible to determine lectin concentrations that produce interpretable data without inducing significant cytotoxicity even when using apoptogenic lectins. That apoptosis is associated with a marked decrease in cell surface sialylation was confirmed by using the sialic acid-specific lectins Maackia amurensis agglutinin and Sambucus nigra agglutinin. These observations were corroborated by lectin blotting analysis with the same lectins., Conclusions: Species- and cell-dependent altered glycosylation patterns are likely to be associated with different modes of apoptosis. The easy and versatile method described in this report should be useful for exploring this field., (2004 Wiley-Liss, Inc.)