Prognostic impact of lymph node metastasis in distal cholangiocarcinoma.

BACKGROUND: The aim of the study was to investigate the prognostic impact of lymph node metastasis in cholangiocarcinoma using three different classifications. METHODS: Patients who underwent pancreaticoduodenectomy for distal cholangiocarcinoma in 24 hospitals in Japan between 2001 and 2010 were included. Survival was calculated by means of the Kaplan-Meier method and differences between subgroups were assessed with the log rank test. The Cox proportional hazards model was used to identify independent predictors of survival. χ(2) scores were calculated to determine the cut-off value of the number of involved nodes, lymph node ratio (LNR) and total lymph node count (TLNC) for discriminating survival. RESULTS: Some 370 patients were included. The median (range) TLNC was 19 (3-59). Nodal metastasis occurred in 157 patients (42·4 per cent); the median (range) number of involved nodes and LNR were 2 (1-19) and 0·11 (0·02-0·80) respectively. Four or more involved nodes was associated with a significantly shorter median survival (1·3 versus 2·2 years; P = 0·001), as was a LNR of at least 0·17 (1·4 versus 2·3 years; P = 0·002). Involvement of nodes along the common hepatic artery, present in 21 patients (13·4 per cent), was also associated with a shorter survival (median 1·3 versus 2·1 years; P = 0·046). Multivariable analysis among 157 node-positive patients identified the number of involved nodes as an independent prognostic factor (risk ratio 1·87; P = 0·002). CONCLUSION: The number of involved nodes was a strong predictor of survival in patients with distal cholangiocarcinoma.

Preoperative MRI for predicting pathological changes associated with surgical difficulty during laparoscopic cholecystectomy for acute cholecystitis.

BACKGROUND: Severe inflammation with necrosis and fibrosis of the gallbladder in acute cholecystitis increases operative difficulty during laparoscopic cholecystectomy. This study aimed to assess the use of preoperative MRI in predicting pathological changes of the gallbladder associated with surgical difficulty. METHODS: Patients who underwent both preoperative MRI and early cholecystectomy for acute cholecystitis between 2012 and 2018 were identified retrospectively. On the basis of the layered pattern of the gallbladder wall on MRI, patients were classified into three groups: high signal intensity (HSI), intermediate signal intensity (ISI), and low signal intensity (LSI). The endpoint was the presence of pathological changes of the gallbladder associated with surgical difficulty, such as necrosis, abscess formation and fibrosis. RESULTS: Of 229 eligible patients, pathological changes associated with surgical difficulty were found in 17 (27 per cent) of 62 patients in the HSI group, 84 (85 per cent) of 99 patients in the ISI group, and 66 (97 per cent) of 68 patients in the LSI group (P < 0·001). For detecting these changes, intermediate to low signal intensity of the gallbladder wall had a sensitivity of 90 (95 per cent c.i. 84 to 94) per cent, specificity of 73 (60 to 83) per cent and accuracy of 85 (80 to 90) per cent. CONCLUSION: Preoperative MRI predicted pathological changes associated with surgical difficulty during laparoscopic cholecystectomy for acute cholecystitis.

ANTECEDENTES: La inflamación grave con necrosis y fibrosis de la vesícula biliar en la colecistitis aguda aumenta la dificultad quirúrgica durante la colecistectomía laparoscópica. Este estudio tuvo como objetivo evaluar el uso de la resonancia magnética preoperatoria (magnetic resonance imaging, MRI) para predecir los cambios patológicos de la vesícula biliar asociados con la dificultad quirúrgica. MÉTODOS: Los pacientes que se sometieron tanto a MRI preoperatoria como a colecistectomía precoz por colecistitis aguda entre 2012 y 2018 fueron identificados retrospectivamente. En base a la distribución en capas de la pared de la vesícula biliar en la MRI, los pacientes se clasificaron en tres grupos: (1) intensidad de señal alta (high signal intensity, HSI), (2) intensidad de señal intermedia (intermediate signal intensity, ISI) y (3) intensidad de señal baja (low signal intensity, LSI). El objetivo final fue la presencia de cambios patológicos en la vesícula biliar asociados con la dificultad quirúrgica, tales como necrosis, formación de abscesos y fibrosis. RESULTADOS: De los 229 pacientes elegibles, se documentaron cambios patológicos asociados con dificultad quirúrgica en 17 (27,4%) de 62 pacientes en el grupo HSI, 84 (84,8%) de 99 pacientes en el grupo ISI y 66 (97,1%) de 68 pacientes en el grupo LSI (P < 0,001). Para detectar estos cambios, la intensidad de señal de intermedia a baja de la pared de la vesícula biliar tuvo una sensibilidad del 89,8% (i.c. del 95% 84,2%-94,0%), una especificidad del 72.6% (i.c. del 95% 59,8%-83,1%) y una precisión del 85,2% (i.c. del 95% 79,9%-89,5%). CONCLUSIÓN: La MRI preoperatoria predijo los cambios patológicos asociados con la dificultad quirúrgica durante la colecistectomía laparoscópica por colecistitis aguda.

Randomized clinical trial of single skin sterilization with a povidone-iodine applicator versus conventional skin sterilization in abdominal surgery.

Background: The efficacy of widely used povidone-iodine applicators for skin sterilization in abdominal surgery is unclear. The aim of this trial was to evaluate whether sterilization with a povidone-iodine applicator was not inferior to a conventional sterilization method. Methods: Patients undergoing elective abdominal surgery were assigned randomly to receive single sterilization with the applicator or conventional sterilization. The primary endpoint was wound infection rate. Secondary endpoints were rate of organ/space surgical-site infection (SSI), adverse effects of povidone-iodine, amount of povidone-iodine used and total cost of sterilization. Results: Of 498 patients eligible for the study between April 2015 and September 2017, 240 were assigned and analysed in the applicator group and 246 in the conventional group. Wound infection was detected in 16 patients (6·7 per cent) in the applicator group and 16 (6·5 per cent) in the conventional group (absolute difference 0·0016 (90 per cent c.i. -0·037 to 0·040) per cent; P = 0·014 for non-inferiority). There was no difference between the groups in the organ/space SSI rate (11 patients (4·6 per cent) in the applicator group and 16 (6·5 per cent) in the conventional group. Both the amount of povidone-iodine used and the total cost of sterilization were higher in the conventional group than in the applicator group (median 76·7 versus 25 ml respectively, P < 0·001; median €7·0 versus €6·4, P < 0·001). Skin irritation was detected in three patients in the conventional group. Conclusion: In abdominal surgery, this povidone-iodine applicator was not inferior to conventional sterilization in terms of the wound infection rate, and it is cheaper. Registration number: UMIN000018231 (http://www.umin.ac.jp/ctr/).

Comparison between dexmedetomidine and midazolam as a sedation agent with local anesthesia in inguinal hernia repair: randomized controlled trial.

PURPOSE: In Japan, inguinal hernia repair is widely performed with local anesthesia. The objective of this study was to evaluate safety and efficacy of intravenous dexmedetomidine as a sedation agent with local anesthesia in inguinal hernia repair. METHODS: We performed this randomized, single-blind study for 200 patients who were scheduled to undergo inguinal hernia repair with local anesthesia. Patients were randomly divided into two groups (dexmedetomidine group: Group D, midazolam group: Group M). The primary outcome was to evaluate the safety of intravenous dexmedetomidine. Secondary outcomes were to analyze results of operators' surveys and patients' questionnaires and evaluate implementation of conscious sedation. RESULTS: Incidence of respiratory depression was significantly higher in Group M than Group D (p = 0.03). Other adverse events examined did not differ significantly. All three operators' questionnaires indicated that results were better in Group D than Group M. More than 70% of patients in both groups were satisfied with the surgery. More than 80% of Group D patients and 74% of Group M patients achieved a state of conscious sedation. CONCLUSION: This study demonstrated that intravenous dexmedetomidine during hernia repair with local anesthesia is safe and the results were satisfactory to both operators and patients.

Small-angle X-ray scattering and computer-aided molecular modeling studies of 20 kDa fragment of porcine amelogenin: does amelogenin adopt an elongated bundle structure?

Amelogenins, which are major matrix constituents in the developing tooth, play a regulatory role in the process of enamel crystal formation. Porcine amelogenin with 173 amino acid residues is rich in proline, glutamine, leucine, and histidine. We utilized the small-angle X-ray scattering (SAXS) technique to examine the solution structure of porcine amelogenin. Samples used were two porcine amelogenins with apparent molecular weights of 20 kDa (amino acids 1 to 148) and 13 kDa (amino acids 46 to 148) on SDS-PAGE. Prior to SAXS measurements, the protein samples were dissolved in 2% (v/v) acetic acid to give a concentration range up to 10 mg/ml. Comparison between Rg (the overall radius of gyration) and Rc (the cross-sectional radius of gyration) revealed that the 20 kDa amelogenin exists in this solution as asymmetric particles with a length of about 15 nm, presumably corresponding of dimers. Based on these experimental data and computer-aided molecular modeling studies, we propose that the 20 kDa amelogenin adopts an elongated bundle structure which mainly consists of extended structures similar to polyproline II and/or beta-strand, interspersed with beta-turn or loop.

Molecular conformation of porcine amelogenin in solution: three folding units at the N-terminal, central, and C-terminal regions.

Circular dichroism (CD) studies were conducted to gain a better insight into the conformation of amelogenins, which were isolated from developing enamel of piglets. The intact porcine amelogenin and its degraded products were purified chromatographically. The 25-residue peptide corresponding to the segment at the C-terminus was synthesized. CD spectra of these samples were measured at pH 5.0-5.3 in the temperature range between 4 and 90 degrees C. The most remarkable finding was that the CD spectrum of the intact amelogenin was accounted for by the sum of the spectra of the three fragments at the N-terminal, central, and C-terminal regions, supporting the hypothesis that the structure of the whole protein consists of discrete folding units. Furthermore, low-angle laser light scattering analysis provided evidence that the 20 kDa amelogenin, the most abundant extracellular matrix protein in forming enamel tissue, exists in a monomeric form at pH 5.3 and 25 degrees C. It was tentatively concluded that the N-terminal region contains beta-sheet structures, while the spectral characteristics of the C-terminal region are similar to those of a random coil conformation. The conformation of the central region was characterized by a strong negative ellipticity at 203 nm, although its nature remains to be defined.

Computer-assisted reconstruction of enamel fissures and carious lesions of human premolars.

The origin of cariogenesis in occlusal fissures remains elusive because of limited information about both the fissure structure and the morphologic details of carious lesions occurring there. The present study was aimed at reconstructing the coronal structure of human premolars with the aid of computer-assisted image analysis and, on the basis of the reconstructed structures, investigating the configuration of fissures, their proximity to the enamel-dentin junction (EDJ), and the occurrence and extension of carious lesions around the fissure walls. The coronal portions of the teeth were embedded in polyester resin and then ground off stepwise by hand. Serial images of the enamel contour on successive ground planes were recorded by objective microscopy after it was distinctly demarcated with either 0.5 w/v% carbol fuchsin or 1 w/v% basic fuchsin. Usually, from 150 to 250 photoprints were collected from a single enamel crown to reproduce the details of the fissure structures. The enamel outline (the occlusal and lateral surfaces, fissure walls, and EDJ) and, if present, the outline of caries-attacked areas were traced accurately on those photoprints by means of a digitizing system. The enamel contour data in series were input into an image analyzing system (Nikon COSMOZONE, 2SA). Observation of the computer-drawn coronal enamel allowed us to view the morphologic features of the fissures (shape, divarication, and proximity to EDJ) and to scrutinize the pathway of enamel caries along fissure walls.(ABSTRACT TRUNCATED AT 250 WORDS)

Preparation of hydroxyapatite crystals and their behavior as seeds for crystal growth.

Samples of crystalline hydroxyapatite [Ca5OH(PO4)3 HA] were prepared by precipitation from aqueous media under a variety of experimental conditions (temperature, concentration of reagents, rates of addition of the reagents, and seeding). The resultant products showed a wide range of particle sizes, i.e., specific surface areas, from 6.39 to 50.1 m2/g. In these preparations, relatively large crystals were obtained with low rates of addition of the reagents or by seeding the precipitating medium. Small differences in super-saturation of the reaction medium can markedly affect the particle sizes and crystalline habits of the resulting products, possibly by altering the processes of nucleation and subsequent crystal growth. When these crystalline materials were used as seeds to study the crystal growth of HA, it was confirmed that the precipitation rate of calcium apatite on seed crystals is highly dependent on the surface areas available for growth, rather than on the particle sizes and amounts of the seed crystals. Small differences in the kinetic runs were observed between the various seed crystals, which can be attributed to differences in the surface properties of these crystals. Additionally, transmission electron microscopy of the seed crystals revealed that some projections form, possibly on the basal planes of the crystals, during crystal growth. Since the growth rate of these projections was greater than the mean growth rate calculated on the basis of changes in solution composition and total surface area, it appears that the kinetics of the growth process is determined, to some extent, by the geometry of the seeds.

Reversible structural changes of octacalcium phosphate and labile acid phosphate.

Acid phosphate is one of the major impurities incorporated into bioapatites, and its quantity and environment in forming mineral have been used as diagnostic probes to pursue acidic precursor(s). Currently, little is known about the structural feature of nonstoichiometric octacalcium phosphate (OCP), which has been advocated to be, most plausibly, mineral salt initially formed during amelogenesis. In the present report, we attempt to define the state of acid phosphate in OCP crystals which were Ca-deficient and contained 40% total phosphate as acid phosphate. We assessed fractions of acid phosphate in discrete environments by extracting the crystals in either deionized water, 10 mmol/L NaOH solution (initial pH 11), or 150 mmol/L Tris buffer at pH 7.4. Solid samples before and after the treatments were examined by chemical analyses and x-ray diffraction. The results indicated that successive extractions with use of the alkaline solution brought about a reversible change (not hydrolysis) in the interior structure of OCP, which accompanied a marked decrease in acid phosphate. A substantial part of the lost acid phosphate was restored during subsequent treatments at neutral pH, and, intriguingly, this restoration accompanied a re-ordering of OCP structure. The data suggested that the acid phosphate in OCP is separated into three pools: (a) a stable pool corresponding to roughly 50 to 60% of the total acid phosphate, (b) a reversibly exchangeable pool corresponding to 25 to 30% of the acid phosphate which may exist either in the water layer or on crystal surfaces, and (c) an unstable (or irreversibly lost) pool corresponding to 15 to 20% of the acid phosphate, a part of which might be explained by the presence of excess hydrogen in OCP. The present work supports the concept that protons and, to a lesser magnitude, phosphate species can diffuse into and out of the OCP lattice prior to initiation of its hydrolytic transition into apatite.

Changes in acid-phosphate content in enamel mineral during porcine amelogenesis.

The present study was undertaken to investigate changes in the acid-phosphate content of porcine enamel mineral during its development and to assess separately the HPO4(2-) pools in labile and stable forms. Enamel samples at the secretory and maturing stages of amelogenesis were obtained from the permanent incisors of five- to six-month-old slaughtered piglets. Human enamel from erupted, extracted teeth, synthetic hydroxyapatite, and carbonatoapatite containing acid phosphate were included as references. The acid-phosphate content of each sample was determined chemically through its pyrolytic conversion to pyrophosphate. The assessment of HPO4(2-) in labile forms was made by analysis of samples preequilibrated with solutions containing 3 mmol/L phosphate at pH11 (to de-protonate the HPO4(2-) species on crystal surfaces). The analytical results of porcine enamel samples showed that: (a) the outermost secretory (youngest) enamel contained the highest HPO4(2-), corresponding to about 16% of the total phosphate; (b) the acid-phosphate content decreased gradually to 10% in the inner (older) secretory and to 6% in the maturing tissue; (c) a substantial part of the HPO4(2-) in developing enamel tissue (50-60% of the HPO4(2-) for the secretory enamel) was in labile forms; and (d) the pool of the labile HPO4(2-) decreased with the growth of enamel mineral. In parallel studies with mature human enamel, it was ascertained that the total acid phosphate was only about 3% of the total phosphate, much lower than in developing porcine enamel, and that the labile pool of HPO4(2-) was also small, corresponding to about 15% of the total acid phosphate determined.(ABSTRACT TRUNCATED AT 250 WORDS)

Possible function of matrix proteins in fluoride incorporation into enamel mineral during porcine amelogenesis.

The present study was undertaken to elucidate the mechanism of fluoride incorporation into secretory enamel mineral, with porcine enamel used as a model. Although the fluoride content in the enamel varied greatly among the animals, we observed that the fluoride-to-calcium ratio in the enamel tissue was maximal at the beginning of the secretory stage; the F/Ca ratio decreased (and leveled off) with the advancement of mineralization. In vitro work showed that some of the fluoride in the secretory enamel tissue was removed with the extraction of organic matter, mostly amelogenins. Furthermore, coating hydroxyapatite crystals with enamel matrix proteins resulted in a retardation of fluoride incorporation into the crystals when exposed to fluoride solutions, as a result of an inhibition of apatite reprecipitation. We also confirmed that the growth kinetics of fluoridated apatite onto HA seeds decreased with increasing coverage of the seed surface with the enamel proteins. All the results of the present study strongly suggest that the fluoride incorporation into enamel mineral during the secretory stage may be regulated by the kinetics of mineralization, which is highly dependent on the driving force for precipitation and the presence of proteinaceous inhibitors, mainly amelogenins.

Labile or surface pools of magnesium, sodium, and potassium in developing porcine enamel mineral.

The present study was undertaken to assess the labile or surface pools of Mg, Na, and K ions in porcine enamel tissues at various developmental stages. The enamel samples, corresponding to the outer and the inner secretory, the early maturing, and the mature hard enamel, were dissected from the labial sides of permanent incisors of 6- to 8-month-old piglets. Each enamel sample was extracted successively with solutions of de-ionized water and 50 mmol/L Tris-4 mol/L guanidine buffer (for removal of organic matrix proteins, mainly amelogenins). The labile (free or organically bound) pools of Mg, Na, and K were assessed by the total amounts of these ionic species extracted by the water and Tris-guanidine buffer. The surface (adsorbed onto enamel mineral) pool of Mg was assessed directly by determination of the adsorption of Mg onto enamel mineral at various developmental stages. The results showed that: (i) 30-40% of the Mg in the secretory and early maturation enamel was in the surface pool (adsorbed onto the enamel mineral); (ii) 25 to 40% of the total sodium in the enamel samples was in labile forms; and (iii) most (around 70-80%) of the total potassium was readily extracted in water and appeared to originate from the enamel fluid; only marginal portions remained in the solids. The present adsorption studies also indicated that the maximum uptake of magnesium in the early maturation enamel was due mostly to an increase of the occupancy by Mg ions of adsorption sites on the crystal surfaces, which become accessible with a massive removal of enamel matrix proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

Proteins in the enamel fluid of immature porcine teeth.

The fluid was separated from the immature soft enamel of porcine permanent teeth in the secretory stage according to procedures reported previously (Aoba and Moreno, 1987). The protein content of the fluid was about 2.8% w/v; its amino-acid composition was characterized by high contents of Pro, Glx, Leu, and His, showing composition similar to that of the 20 kilo-dalton (kd) amelogenin or its C-terminal segments. The two major protein species in the fluid had apparent molecular weights of 13 kd and 11 kd, as determined by SDS electrophoresis; the N-terminal residue of the former was Leu, while that of the latter was Ala. The C-terminal sequence of both of them was -Met-Phe-Ser. By comparison with the published sequence of 20-kd porcine amelogenin, it is concluded that the main fluid constituents were derived by cleavages of N-terminal segments from the 20-kd amelogenin.

Effect of fluoride in the apatitic lattice on adsorption of enamel proteins onto calcium apatites.

The selective adsorption of enamel proteins onto crystalline calcium apatites having different specific surface areas and various degrees of fluoride substitution was investigated. The proteins were obtained from the outer (close to the ameloblast) layer of secretory enamel of porcine permanent incisors. The adsorption of the enamel proteins was not affected markedly by the variation of specific surface area of the hydroxyapatites used as adsorbents, but it was enhanced substantially with increasing fluoride content in the crystalline lattice. Through the use of SDS- and two-dimensional polyacrylamide gel electrophoresis, it was shown that the originally secreted amelogenin (25 kd) as well as 60-90-kd and 5-6-kd molecules adsorbed most selectively onto the hydroxyapatites and that additional moieties having 21-23-kd and 14-18-kd molecular masses commenced to adsorb onto the apatitic surfaces with increasing degrees of fluoride substitution in the lattice. In contrast, the 20-kd amelogenin, a product partially degraded from the 25-kd amelogenin, showed no significant adsorption, even onto the fluoridated apatites. These results suggest that the retention of proteinaceous matrix in the developing enamel might be affected by the nature of the forming crystals.

High-resolution electron microscopy of hydroxyapatite grown in dilute solutions.

High-resolution electron microscopy (HREM) was applied to the study of seeded crystal growth of hydroxyapatite (HA) in supersaturated solutions. The HA seed crystals were rod-shaped, elongated along the c-axis, and showed smooth contours. The seed crystals exhibited hexagonal prism facets, and one end was rhombohedrally terminated, whereas the opposite end was blunt. Growth experiments were carried out at 37 degrees C, and solution compositions were carefully selected to avoid the involvement of precursor phases during the HA growth. After five-hour growth, the total amount of precipitation was from 3 to 7% of the initial crystal mass added to the solutions; examination of the crystals by HREM disclosed the formation of projections on the end surfaces of the HA crystals. High-resolution TEM clearly showed lattice fringes with predictable spacings over the entire crystal specimen, including the formed projections. Analysis of selected regions by optical diffraction of the HRTEM fringe negatives showed that the nature and orientation of the projections were similar to those of the underlying seed crystal. High-resolution SEM of the HA crystal after five-hour growth disclosed step-like structures on the prism faces, as well as the projections localized on the ends. These results strongly suggest that more than one growth process may be involved, perhaps related to the distinct faces of the HA seed crystals.

Orientations of carbonate ions in human tooth enamel studied with use of the CO3-3 radical ions as probes.

With the use of CO3-3 radical ions as probes, it is concluded that the carbonate ions substitute in the apatite lattice of human tooth enamel in two different ways. Most of the carbonate ions in the apatite lattice are aligned with their symmetry axis parallel to the c axis, whereas the remaining carbonate ions are aligned with their symmetry axis perpendicular to the c axis.

Organic-bound chromium in enamel, dentin, and bone studied by electron spin resonance.

The present ESR investigation shows clearly that chromium is potentially present in the calcified tissues. With the use of the hydrazine-deproteinated procedure, it is suggested that chromium is associated with the organic constituent of the calcified tissues. The fact that the unheated tissues disclosed no chromium signal indicates either that the potentially present chromium is not paramagnetic, or that it is still paramagnetic, but the chromium signal cannot be detected because of a short relaxation time due to strong interaction with the organic materials.

Dissolution rate behavior of fluoridated apatite pellets.

The dissolution rate of synthetic fluoridated apatite pellets with various fluoride content was measured in acetate buffer solution at pH 4.0. The dissolution rate decreased with degree of fluoridation. The rate decreased greatly at low fluoride levels. The experimental data were analyzed by means of a diffusion-controlled model and compared with the solubility data. This model showed that the initial dissolution rate is greatly affected by both the solubility and the overall mass transfer resistance. Overall mass transfer resistance is affected by fluoride content.

The laser-Raman spectroscopic studies on human enamel and precipitated carbonate-containing apatites.

Laser-Raman spectra of synthetic apatites containing from zero to 12.4 wt% carbonate ions and enamel apatite are reported. Special attention is paid to the correlation between the intensities of the carbonate, phosphate, and hydroxyl modes and the carbonate content in the apatites. The data presented here indicate that most of the CO2-3 incorporated in the apatite lattice substitutes for PO3-4 and suggest that a small amount of CO2-3 might be located in the OH- position.

Solubility and crystallinity in relation to fluoride content of fluoridated hydroxyapatites.

Fluoridated hydroxyapatites were synthesized at 80 degrees C (pH 7.4) to study their crystallinity and solubility in relation to fluoride content. The crystallinity of fluoridated hydroxyapatities increased initially, then decreased, and finally increased again with the degree of fluoridation, whereas the decrease in relative solubility of fluoridated hydroxyapatites at 25 degrees C (pH 4.0) was proportional to the logarithm of the fluoride content.