Assessment of the intrasinusidal volume before and after maxillary sinus augmentation using mri - a pilot study of eight patients.

PURPOSE: The purpose of this study was to evaluate the suitability, accuracy, and reliability of a non-invasive 3-Tesla magnetic resonance imaging technique (3 T-MRI) for the visualization of maxillary sinus grafts in comparison to conventional, X-ray-based, established standard imaging techniques. METHODS: A total of eight patients with alveolar bone atrophy who required surgical sinus floor augmentation in the course of dental implantation were included in this pilot study. Alongside pre-operative cone-beam computed tomography (CBCT), 3 T-MRI was performed before and 6 months after sinus floor augmentation. Two investigators measured the maxillary sinus volume preoperatively and after bone augmentation. RESULTS: In all cases, MRI demonstrated accurately the volumes of the maxillary sinus grafts. Following surgery, the bony structures suitable for an implant placement increased at an average of 4.89 cm3, corresponding with the decrease of the intrasinusidal volumes. In general, interexaminer discrepancies were low and without statistical significance. CONCLUSION: In this preliminary study, we could demonstrate the feasibility of MRI bone volume measurement as a radiation-free alternative with comparable accuracy to CT/CBCT before procedures like sinus floor augmentation. Nevertheless, costs and artifacts, also present in MRI, have to be taken into account. Larger studies will be necessary to justify the practicability of MRI bone volume evaluation.

Routine Gastric Decompression after Pancreatoduodenectomy: Treating the Surgeon?

BACKGROUND: The decision to routinely leave a nasogastric tube after pancreatoduodenectomy remains controversial. We sought to determine the impact of immediate nasogastric tube removal versus early nasogastric tube removal (<24 h) on postoperative outcomes. METHODS: A retrospective review of our institution's prospective ACS-NSQIP database identified patients that underwent pancreatoduodenectomy from 2015 to 2018. Outcomes were compared among patients with immediate nasogastric tube removal versus early nasogastric tube removal. RESULTS: A total of 365 patients were included in primary analysis (no nasogastric tube, n = 99; nasogastric tube removed <24 h, n = 266). Thirty-day mortality and infectious, renal, cardiovascular, and pulmonary morbidity were similar in comparing those with no nasogastric tube versus early nasogastric tube removal on univariable and multivariable analyses (P > 0.05). Incidence of delayed gastric emptying (11.1 versus 13.2%) was similar between groups. Patients with no nasogastric tube less frequently required nasogastric tube reinsertion (n = 4, 4%) compared to patients with NGT <24 h (n = 39, 15%) (OR = 3.83, 95% CI [1.39-10.58]; P = 0.009). CONCLUSION: Routine gastric decompression can be safely avoided after uneventful pancreaticoduodenectomy.

Optimal hepatobiliary scintigraphy for gallbladder dyskinesia.

BACKGROUND: The accuracy of hepatobiliary scintigraphy to assess gallbladder function remains controversial. National supply shortages of pharmaceutical-grade cholecystokinin led to the use of an oral fatty meal to stimulate gallbladder contraction during hepatobiliary scintigraphy. The goal of this study was to compare the predictive indices of cholecystokinin and fatty meal ingestion for stimulation of gallbladder contraction. METHODS: Patients evaluated with hepatobiliary iminodiacetic acid scan from 2014 to 2017 were reviewed and grouped based on testing stimulant (fatty meal versus cholecystokinin). Patients who later underwent cholecystectomy were selected for analysis. Hepatobiliary iminodiacetic acid results were correlated with surgical pathology and postoperative resolution of symptoms. Two-way statistical analysis was performed. RESULTS: A total of 359 patients underwent hepatobiliary iminodiacetic acid scan followed by cholecystectomy for biliary dyskinesia. Patients who received fatty meal stimulant (n = 86) were compared to those that received cholecystokinin (n = 273). Mean gallbladder ejection fraction during hepatobiliary iminodiacetic acid was 38% and 44% for the cholecystokinin and fatty meal groups, respectively, P = .073. Predictive metrics were not statistically different between groups with regard to pathology, symptomatic improvement, or accuracy. Symptomatic resolution (cholecystokinin-hepatobiliary iminodiacetic acid 78%, fatty meal-hepatobiliary iminodiacetic acid 68%; P = 0.058) and specificity (cholecystokinin-hepatobiliary iminodiacetic acid 26%, fatty meal-hepatobiliary iminodiacetic acid 44%, P = 0.417) were comparable in both testing groups. CONCLUSION: Stimulation of gallbladder contraction with a fatty meal during hepatobiliary iminodiacetic acid testing is a more affordable and reliable alternative to cholecystokinin for patients undergoing evaluation for gallbladder dysmotility.

Hepatic Steatosis After Neoadjuvant Chemotherapy for Pancreatic Cancer: Incidence and Implications for Outcomes After Pancreatoduodenectomy.

BACKGROUND: This study aimed to determine the incidence of new onset hepatic steatosis after neoadjuvant chemotherapy for pancreatic cancer and its impact on outcomes after pancreatoduodenectomy. METHODS: Retrospective review identified patients who received neoadjuvant chemotherapy for pancreatic adenocarcinoma and underwent pancreatoduodenectomy from 2013 to 2018. Preoperative computed tomography scans were evaluated for the development of hepatic steatosis after neoadjuvant chemotherapy. Hypoattenuation included liver attenuation greater than or equal to 10 Hounsfield units less than tissue density of spleen on noncontrast computed tomography and greater than or equal to 20 Hounsfield units less on contrast-enhanced computed tomography. RESULTS: One hundred forty-nine patients received neoadjuvant chemotherapy for a median of 5 cycles (interquartile range (IQR), 4-6). FOLFIRINOX was the regimen in 78% of patients. Hepatic steatosis developed in 36 (24%) patients. The median time from neoadjuvant chemotherapy completion to pancreatoduodenectomy was 40 days (IQR, 29-51). Preoperative biliary stenting was performed in 126 (86%) patients. Neoadjuvant radiotherapy was delivered to 23 (15%) patients. Female gender, obesity, and prolonged exposure to chemotherapy were identified as risk factors for chemotherapy-associated hepatic steatosis. Compared with control patients without neoadjuvant chemotherapy-associated hepatic steatosis, patients developing steatosis had similar rates of postoperative pancreatic fistula (8% (control) vs. 4%, p = 0.3), delayed gastric emptying (8% vs. 14%, p = 0.4), and major morbidity (11% vs. 15%, p = 0.6). Ninety-day mortality was similar between groups (8% vs. 2%, p = 0.08). CONCLUSION: Hepatic steatosis developed in 24% of patients who received neoadjuvant chemotherapy but was not associated with increased morbidity or mortality after pancreatoduodenectomy.

Comparison of skin closure techniques in patients undergoing open pancreaticoduodenectomy: A single center experience.

BACKGROUND: This study evaluated closure techniques and incisional surgical site complications (SSCs) and incisional surgical site infections (SSIs) after pancreaticoduodenectomy (PD). METHODS: Retrospective review of open PDs from 2015 to 2018 was performed. Outcomes were compared among closure techniques (subcuticular + topical skin adhesive (TSA); staples; subcuticular only). SSCs were defined as abscess, cellulitis, seroma, or fat necrosis. SSIs were defined according to the National Surgical Quality Improvement Program (NSQIP). RESULTS: Patients with subcuticular + TSA (n = 205) were less likely to develop an incisional SSC (9.8%) compared to staples (n = 139) (20.1%) and subcuticular (n = 74) (16.2%) on univariable analysis (P = 0.024). Multivariable analysis revealed no statistically significant difference in incisional SSC between subcuticular + TSA and subcuticular (P = 0.528); a significant difference remained between subcuticular + TSA and staples (P = 0.014). Unadjusted median length of stay (LOS) (days) was significantly longer for staples (9) vs. subcuticular (8) vs. subcuticular + TSA (7); P < 0.001. Incisional SSIs were evaluated separately according to the NSQIP definition. When comparing rates, the subcuticular + TSA group experienced lower incisional SSIs compared to the other two techniques (4.9% vs. 10.1%, 10.8%). However, this difference was not statistically significant by either univariable or multivariable analysis. CONCLUSIONS: Subcuticular suture + TSA reduces the risk of incisional SSCs when compared to staples alone after pancreaticoduodenectomy.

Comparison of human turning gait with the mechanical performance of lower limb prosthetic transverse rotation adapters.

Given the importance of minimizing transverse plane shear stress on soft tissue, several transverse rotational adapters (TRAs) are available for incorporation in lower limb prostheses. This study compares kinetic and kinematic data from human subjects during straight and turning gaits to the mechanical performance of several TRAs. Physiological data were collected from three individuals walking straight and turning at self-selected speeds around a 1 m radius circle. The average peak torques and range of motion for normal subjects while turning were 8.2 Nm and 26 degrees (outside leg), 11.8Nm and 20 degrees (inside leg), and 11.4 Nm and 20 degrees (right leg) during straight gait. Each TRA was mechanically tested without axial loading in a servo-hydraulic material testing system (MTS) over its rotational range at 0.5 dergrees/s and 60 degrees/s. The TRAs with axial compression were also tested at 0.5 degrees/s under a 736N (75kg mass) axial load. Applying these torques to the different TRAs yielded 3 to 35 degrees rotation, depending on the elastomer installed. Some TRAs had nearly constant stiffness, while others stiffened with rotation. The TRAs also varied in their average maximum stiffness from 0.4Nm/degree to 2.7Nm/degrees. Normal subjects exhibit interior vs. exterior asymmetrical torques and displacements; however, only one of the TRAs is designed to allow asymmetrical stiffness, and none have asymmetric ranges. Prosthetists and physicians can use these data to better interpret amputees' qualitative remarks and to prescribe the correct TRA and/or elastomer. This information also forms a basis for further design and development of novel torque absorbing prosthetic adapters.

Role of nonimmune IgG bound to PfEMP1 in placental malaria.

Infections with Plasmodium falciparum during pregnancy lead to the accumulation of parasitized red blood cells (infected erythrocytes, IEs) in the placenta. IEs of P. falciparum isolates that infect the human placenta were found to bind immunoglobulin G (IgG). A strain of P. falciparum cloned for IgG binding adhered massively to placental syncytiotrophoblasts in a pattern similar to that of natural infections. Adherence was inhibited by IgG-binding proteins, but not by glycosaminoglycans or enzymatic digestion of chondroitin sulfate A or hyaluronic acid. Normal, nonimmune IgG that is bound to a duffy binding-like domain beta of the P. falciparum erythrocyte membrane protein 1 (PfEMP1) might at the IE surface act as a bridge to neonatal Fc receptors of the placenta.

Cloning, sequence and crystallographic structure of recombinant iron superoxide dismutase from Pseudomonas ovalis.

The gene encoding the iron-dependent superoxide dismutase from Pseudomonas ovalis was cloned from a genomic library and sequenced. The ORF differs from the previously published protein sequence, which was used for the original structure determination, at 16 positions. The differences include three additional inserted residues, one deleted residue and 12 point substitutions. The gene was subcloned and the recombinant protein overexpressed, purified and crystallized in a trigonal space group. The structure was determined by molecular replacement and was refined to 2.1 A resolution.

Regulation of transcription by ubiquitination without proteolysis: Cdc34/SCF(Met30)-mediated inactivation of the transcription factor Met4.

Polyubiquitination of proteins by Cdc34/SCF complexes targets them for degradation by the 26S proteasome. The essential F-box protein Met30 is the substrate recognition subunit of the ubiquitin ligase SCF(Met30). The critical target of SCF(Met30) is the transcription factor Met4, as deletion of MET4 suppresses the lethality of met30 mutants. Surprisingly, Met4 is a relatively stable protein and its abundance is not influenced by Met30. However, transcriptional repression of Met4 target genes correlates with Cdc34/SCF(Met30)-dependent ubiquitination of Met4. Functionally, ubiquitinated Met4 associates with target promoters but fails to form functional transcription complexes. Our data reveal a novel proteolysis-independent function for Cdc34/SCF and indicate that ubiquitination of transcription factors can be utilized to directly regulate their activities.

A novel endonuclease mechanism directly visualized for I-PpoI.

A novel mechanism of DNA endonucleolytic cleavage has been visualized for the homing endonuclease I-PpoI by trapping the uncleaved enzyme-substrate complex and comparing it to the previously visualized product complex. This enzyme employs a unique single metal mechanism. A magnesium ion is coordinated by an asparagine residue and two DNA oxygen atoms and stabilizes the phosphoanion transition state and the 3'oxygen leaving group. A hydrolytic water molecule is activated by a histidine residue for an in-line attack on the scissile phosphate. A strained enzyme-substrate-metal complex is formed before cleavage, then relaxed during the reaction.

DNA polymerase epsilon catalytic domains are dispensable for DNA replication, DNA repair, and cell viability.

DNA polymerase epsilon (Pol epsilon) is believed to play an essential catalytic role during eukaryotic DNA replication and is thought to participate in recombination and DNA repair. That Pol epsilon is essential for progression through S phase and for viability in budding and fission yeasts is a central element of support for that view. We show that the amino-terminal portion of budding yeast Pol epsilon (Pol2) containing all known DNA polymerase and exonuclease motifs is dispensable for DNA replication, DNA repair, and viability. However, the carboxy-terminal portion of Pol2 is both necessary and sufficient for viability. Finally, the viability of cells lacking Pol2 catalytic function does not require intact DNA replication or damage checkpoints.

1,3-Dioxane-4,6-dione-5-carboxamide-based inhibitors of human group IIA phospholipase A: X-ray structure of the complex and interfacial selection of inhibitors from a structural library.

A library of 109 1,3-dioxane-4,6-dione-5-carboxamides was prepared by solution-phase methods as potential inhibitors of human group IIa phospholipase A2. Tight binding inhibitors were found by an interfacial affinity selection method. The crystal structure of the secreted phospholipase A2 containing one of the inhibitors was determined, and it reveals the inhibitor-calcium bidendate coordination.

DNA binding and cleavage by the nuclear intron-encoded homing endonuclease I-PpoI.

Homing endonucleases are a diverse collection of proteins that are encoded by genes with mobile, self-splicing introns. They have also been identified in self-splicing inteins (protein introns). These enzymes promote the movement of the DNA sequences that encode them from one chromosome location to another; they do this by making a site-specific double-strand break at a target site in an allele that lacks the corresponding mobile intron. The target sites recognized by these small endonucleases are generally long (14-44 base pairs). Four families of homing endonucleases have been identified, including the LAGLIDADG, the His-Cys box, the GIY-YIG and the H-N-H endonucleases. The first identified His-Cys box homing endonuclease was I-PpoI from the slime mould Physarum polycephalum. Its gene resides in one of only a few nuclear introns known to exhibit genetic mobility. Here we report the structure of the I-PpoI homing endonuclease bound to homing-site DNA determined to 1.8 A resolution. I-PpoI displays an elongated fold of dimensions 25 x 35 x 80 A, with mixed alpha/beta topology. Each I-PpoI monomer contains three antiparallel beta-sheets flanked by two long alpha-helices and a long carboxy-terminal tail, and is stabilized by two bound zinc ions 15 A apart. The enzyme possesses a new zinc-bound fold and endonuclease active site. The structure has been determined in both uncleaved substrate and cleaved product complexes.

Regulation of cell size by glucose is exerted via repression of the CLN1 promoter.

Yeast cells are keenly sensitive to the availability and quality of nutrients. Addition of glucose to cells growing on a poorer carbon source elicits a cell cycle delay during G1 phase and a concomitant increase in the cell size. The signal is transduced through the RAS-cyclic AMP pathway. Using synchronized populations of G1 cells, we show that the increase in cell size required for budding depends upon CLN1 but not other G1 cyclins. This delay in cell cycle initiation is associated specifically with transcriptional repression of CLN1. CLN2 is not repressed. Repression of CLN1 is not limited to the first cycle following glucose addition but occurs in each cell cycle during growth on glucose. A 106-bp fragment of the CLN1 promoter containing the three MluI cell cycle box (MCB) core elements responsible for the majority of CLN1-associated upstream activation sequence activity is sufficient to confer glucose-induced repression on a heterologous reporter. A mutant CLN2 promoter that is rendered dependent upon its three MCB core elements due to inactivation of its Swi4-dependent cell cycle box (SCB) elements is also repressed by glucose. The response to glucose is partially suppressed by inactivation of SWI4, but not MBP1, which is consistent with the dependence of MCB core elements upon the SCB-binding transcription factor (SBF). We suggest that differential regulation of CLN1 and CLN2 by glucose results from differences in the capacity of SBF to activate transcription driven by SCB and MCB core elements. Finally, we show that transcriptional repression is sufficient to explain the cell cycle delay that occurs in response to glucose.

The wacA gene of Dictyostelium discoideum is a developmentally regulated member of the MIP family.

We isolated a cDNA from Dictyostelium discoideum that encodes a 30 kDa protein with significant similarity to members of the major intrinsic protein (MIP) family of membrane transporters. The most closely related protein in the public data bases is an aquaporin from Cicadella viridis which shows 34% identity. The cDNA was used to isolate and characterize genomic fragments carrying the Dictyostelium gene which we named wacA. Genomic probes were used to recognize wacA mRNA isolated at various stages of development. The results showed that the gene is developmentally regulated such that the mRNA first appears at 12 h of development and is retained throughout the remainder of development. In situ hybridization of whole mounts prepared at 15 h of development showed that wacA mRNA accumulates exclusively in prespore cells and is absent from prestalk cells. Although wacA expression is prespore specific, disruption of the gene by homologous recombination did not result in observable alterations in the formation of spores or their resistance to osmotic challenges.

Crystallization and preliminary X-ray studies of I-PpoI: a nuclear, intron-encoded homing endonuclease from Physarum polycephalum.

The homing endonuclease I-PpoI is encoded by an optional third intron, Pp LSU 3, found in nuclear, extrachromosomal copies of the Physarum polycephalum 26S rRNA gene. This endonuclease promotes the lateral transfer or "homing" of its encoding intron by recognizing and cleaving a partially symmetric, 15 bp homing site in 26S rDNA alleles that lack the Pp LSU 3 intron. The open reading frame encoding I-PpoI has been subcloned, and the endonuclease has been overproduced in E. coli. Purified recombinant I-PpoI has been co-crystallized with a 21 bp homing site DNA duplex. The crystals belong to space group P3(1)21, with unit cell dimensions a = b = 114 A, c = 89 A. The results of initial X-ray diffraction experiments indicate that the asymmetric unit contains an enzyme homodimer and one duplex DNA molecule, and that the unit cell has a specific volume of 3.4 A3/dalton. These experiments also provide strong evidence that I-PpoI contains several bound zinc ions as part of its structure.

Calcineurin-immunosuppressor complexes.

Crystal structures of the Ser/Thr phosphatase calcineurin (protein phosphatase 2B) have recently been solved by X-ray crystallography, both in the free-protein state, and complexed with the immunophilin/immunosuppressant FKBP12/FK506. Core elements of the calcineurin phosphatase have been found to be similar to the corresponding elements of Ser/Thr phosphatase 1 and purple acid phosphatase. The structures provide a basis for understanding calcineurin inhibition by a ternary complex of immunophilin and immunosuppressant proteins.

On the biological role of the nuclear polymerizing NAD+: protein(ADP-ribosyl) transferase (ADPRT): ADPRT from Dictyostelium discoideum and inactivation of the ADPRT gene in the mouse.

Two approaches have been used to elucidate the role of the nuclear polymerizing NAD+:protein(ADP-ribosyl)-transferase (ADPRT): i) comparison of the primary structure of Dictyostelium discoideum ADPRT derived from a 2 kb, partial cDNA sequence with the mammalian, fish, amphibian and insect counterparts revealed an overall homology of 25%. Whereas the automodification domain was not conserved at all, the NAD+ binding domain (aa 859-908) showed more than 70% identical amino acids in all species. Together with the similar enzymatic properties of the ADPRTs the genetic conservation underlined the notion that ADPRT plays a major role in various cellular processes; and ii) inactivation of the ADPRT gene in murine embryonic stem cells by homologous recombination led to mouse strains with a complete lack of nuclear poly(ADP-ribosyl)ation. These ADPRT mutant mice were viable and fertile indicating that ADPRT is dispensable in mouse development. Moreover, repair of UV and MNNG induced DNA damage was not affected in ADPRT/3T3 like fibroblasts, as measured by reactivation of in vitro damaged reporter plasmids and unscheduled DNA synthesis. However, about 30% of the ADPRT mutant mice developed pathological skin aberrations on a mixed 129/Sv x C57B1/6 genetic background. These mice will be extremely useful to define the precise biological role of poly(ADP-ribosyl)ation.