Protein nanoarrays using the annexin A5 two-dimensional crystal on supported lipid bilayers.

Protein nanoarrays are regularly ordered patterns of proteins fixed on a solid surface with a periodicity on the order of nanometers. They have significant potential applications as highly sensitive bioassays and biosensors. While several researchers have demonstrated the fabrication of protein nanoarrays with lithographic techniques and programmed DNA nanostructures, it has been difficult to fabricate a protein nanoarray containing a massive number of proteins on the surface. We now report the fabrication of nanoarrays of streptavidin molecules using a two-dimensional (2D) crystal of annexin A5 as a template on supported lipid bilayers that are widely used as cell membranes. The 2D crystal of annexin A5 has a six-fold symmetry with a period of about 18 nm. There is a hollow of a diameter of about 10 nm in the unit cell, surrounded by six trimers of annexin A5. We found that a hollow accommodates up to three streptavidin molecules with their orientation controlled, and confirmed that the molecules in the hollow maintain their specific binding capability to biotinylated molecules, which demonstrates that the fabricated nanoarray serves as an effective biosensing platform. This methodology can be directly applied to the fabrication of nanoarrays containing a massive number of any other protein molecules.

[A Case of Obstructive Rectal Cancer with Flaccid Constipation Resected One Year and Three Months after SEMS Implantation].

We report a case of rectal cancer that was resected 1 year and 3 months after SEMS implantation. An 89-year-old man was previously diagnosed with sigmoid colon cancer at another hospital but did not undergo surgery. Three years and 7 months after the diagnosis, SEMS was implanted at another hospital. Four years and 10 months after the diagnosis, the patient was diagnosed with intestinal obstruction at our hospital. Since the SEMS was open on colonoscopy, the patient was also suspected of having flaccid constipation. The primary tumor was resected, and a colostomy was constructed in the descending colon. Bridge to surgery for obstructive colorectal cancer was performed within a few weeks after SEMS implantation. At our hospital, resection was performed after a long time. No complications, such as obstruction or perforation, were observed. In addition, although there were concerns regarding increased vascular invasion due to compression and drainage of the cancerous tissue, in our case, the vascular invasion was mild, and no distant metastasis or invasion of other organs was observed. SEMS can be used for long-term implantation and does not necessarily cause cancer progression.

Molecular-Resolution Imaging of Interfacial Solvation of Electrolytes for Lithium-Ion Batteries by Frequency Modulation Atomic Force Microscopy.

Solvation structures formed by ions and solvent molecules at solid/electrolyte interfaces affect the energy storage performance of electrochemical devices, such as lithium-ion batteries. In this study, the molecular-scale solvation structures of an electrolyte, a solution of lithium bis(trifluoromethanesulfonyl)imide (LiTFSI) in propylene carbonate (PC) at the electrolyte-mica interface, were measured using frequency-modulation atomic force microscopy (FM-AFM). The spacing of the characteristic force oscillation in the force versus distance curves increased with increasing ion concentration, suggesting an increase in the effective size of molecules at the interface. Molecular dynamics simulations showed that the effective size of molecular assemblies, namely, solvated ions formed at the interface, increased with increasing ion concentrations, which was consistent with the experimental results. Knowledge of molecular-scale structures of solid/electrolyte interfaces obtained by a combination of FM-AFM and molecular dynamics simulations is important in the design of electrolytes for future energy devices and in improving their properties.

[A Greater Than Four-Year Survival of a Patient with Inoperable Hilar Cholangiocarcinoma following Portal Vein Embolization and Administration of S-1].

We report a patient with inoperable hilar cholangiocarcinoma due to invasion at the umbilical portion who survived more than 4 years after right portal vein embolization and administration of S-1(50 mg/day). A 64-year-old male patient was immediately hospitalized for liver dysfunction and a high level of HbA1c. The disease was diagnosed as hilar cholangiocarcinoma mainly extending along the right hepatic duct. We made a request for operation to Nagoya University. He received right portal vein embolization in order to grow the residual liver but was deemed inoperable because of invasion at the umbilical portion. He refused chemotherapy but accepted administration of S-1(50 mg/day). Approximately 3 months after starting S-1, his ALP level normalized and about 9 months later stenting tube was lost. Subsequently, he returned to his job. Approximately 2 years and 2 months later, administration of S-1 was interrupted due to a harmful side effect. After approximately 13 months without S-1, the levels of CA19-9 and ALP again became elevated and administration of S-1 was restarted. He was temporarily hospitalized for abdominal pain and fever, but quickly recovered. Although CA19-9 and ALP levels re-normalized, he died after returning home. We emphasize the possibility of maintaining long-term health by minimal- dose S-1 therapy for inoperable hilar cholangiocarcinoma.

[Clinical Characteristics, Surgical Outcomes, and Optimal Approach Methods for Transverse Colon Cancers].

Surgery for transverse colon cancer is very difficult because of small number of patients, variations in the vascular system, and complexity of the mobilize hepatic and splenic flexure of colon. We analyzed the clinical characteristics and surgical outcomes in 51 cases who underwent surgery for transverse colon cancer at our hospital between January 2014 and December 2021, and examined the optimal laparoscopic approach method. The surgical procedure was right hemicolectomy in 24 cases, transverse colectomy in 22 cases, and left hemicolectomy in 5 cases, of which 37 cases had laparoscopic surgery. In laparoscopic surgery, when comparing the cranial first approach group in 21 cases and the caudal approach group in 16 cases, the operative time was almost the same between the 2 groups. In the cranial first approach group the amount of bleeding tended to be small, the number of MCA lymph node dissections tended to be large and the hospital stay tended to be short, though there was no statistically significant difference. The results suggest that the cranial first approach tends to be safer and more accurate than the caudal approach.

Surface charge density measurement of a single protein molecule with a controlled orientation by AFM.

The spatial distribution of functional groups causes a charge distribution that often has a close relationship with its biofunctions. To understand them of the protein molecules, measurements of the charge distribution under physiological conditions are desired. Atomic force microscopy (AFM) has been utilized to measure the surface charge density by measuring the electric double layer (EDL) force caused by the overlap of the EDLs on the surfaces of the AFM tip and the biomolecule. Here, we demonstrated the surface charge density measurement of a single streptavidin (SA) protein molecule by the three-dimensional force mapping method based on frequency modulation AFM (FM-AFM). The SA has a strong affinity to biotin because of the electrostatic interactions between the molecules. Therefore, the surface charge density measurements of the biotin-binding sites and other surface areas of the molecule have been anticipated. However, the surface charge density of the surfaces other than the biotin-binding side has never been measured. We demonstrate the surface charge density measurement of the top surface of the single SA molecule, which is perpendicular to the biotin-binding sides, with a controlled orientation using DNA origami as a template by FM-AFM in an electrolyte solution. The surface charge density of the top surface of the SA molecule was estimated by fitting the experimental force curves to the Derjaguin-Landau-Verwey-Overbeck theory. We found that the surface charge density of the top surface of the SA molecule is comparable to those reported earlier for the biotin-binding sides of the molecule. We expect that, by using the DNA origami technology, one can control the orientation of a biomolecule attached to the substrate and measure the surface charge density of the specific surface areas of the biomolecule to obtain information that will help us to understand the relationship between their structures and functions.

[Penrose Method Independent of the Rib Arch for Retraction of the Liver in Laparoscopic Gastrectomy y-Shape+1 Method].

We report the y-shape+1 method, which is a retraction of the liver method, independent of the position of the costal arch. Additionally, we report changes in liver enzymes induced by different methods of liver retraction. Three Penrose No. 6 drains were cut into 6 cm pieces. Penrose 1 was fixed with a thread attached to the end and Penrose 2 in a y-shape. A knitting thread with needles was prepared, and a loop was made at the end of the thread. The needle thread was passed over the abdominal wall and through the Penrose 2. The needle thread was passed over the diaphragmatic leg and through the Penrose 3. The thread was pulled over the abdominal wall and diaphragmatic leg and fixed to the end of Penrose 1 out of the body, and the liver was drained. The rate of change in liver enzymes from the preoperative to postoperative stages was examined separately in the Penrose and Nathanson groups. In the y-shape+1 technique, retraction of the liver can be performed regardless of the position of the rib arch; however, intracorporeal suture ligation is necessary. This method is useful when the conventional Penrose method is inappropriate because of the position of the rib arch.

[Surgical Outcomes and Clinicopathological Analysis of Non-Ampullary Duodenal Tumors].

Non-ampullary duodenal tumors are relatively rare; however, in recent years, they have been encountered more frequently. We analyzed the surgical outcomes and clinicopathological findings in 20 patients who underwent surgery based on preoperative diagnoses of non-ampullary duodenal tumors at our hospital between January 2011 and April 2021. We performed surgery for 3 cases of GIST, 4 cases of adenoma, and 13 cases of adenocarcinoma. The average age of the patients was 64.3 years and the male-to-female ratio was 17:3. The location of the tumor was the blub in 5 cases, the superior duodenal angle in 2 cases, the descending portion in 9 cases, the horizontal portion in 3 cases, and the ascending portion in 1 case. The histological type of adenocarcinoma was tub1 in all cases of early cancer, whereas in advanced cancer, there were many cases with histological types other than tub1. Various surgical procedures from duodenal local resection to pancreatoduodenectomy can be performed for treating non-ampullary duodenal tumors depending on the tumor location and the necessity of lymph node dissection. It is important to establish a treatment policy that considers both curability and invasiveness.

Bochdalek hernia in an adult: two case reports and a review of perioperative cardiopulmonary complications.

BACKGROUND: Bochdalek hernia in an adult is very rare and often needs an immediate surgical repair for the herniation. Although its etiology and surgical techniques have frequently been reported, perioperative complications, especially cardiopulmonary problems, remain unknown. We reported two adults with Bochdalek hernia and reviewed the published literatures with a focus on these issues. CASE PRESENTATION: We experienced two adult cases of Bochdalek hernia with gastrointestinal strangulation. One case had massive herniation of the stomach, colon, spleen, and pancreas in the left chest, causing repeated vomiting. The other had a right-side hernia with strangulation of the colon. We successfully performed emergency repairs of these diaphragmatic hernias without any postoperative complications. CONCLUSIONS: Our literature review revealed that life-threatening cardiopulmonary complications, such as empyema or cardiac arrest caused by the tamponade effect of the herniated viscera, sometimes occurred in patients with Bochdalek hernia. These complications were found in Bochdalek hernia with gastrointestinal strangulation.

[The Efficacy of Endoscopic Esophageal Stent Placement for Malignant Esophageal Stenosis].

Malignant esophageal stenosis adversely affects the quality of life(QOL)on account of poor oral intake. Endoscopic esophageal stenting is one of palliative therapy for improve QOL because of minimally invasive and simple procedure. We investigated the outcomes of esophageal stenting in our institution. Twenty patients with malignant esophageal stenosis who underwent esophageal stenting in our institution between April 2014 and December 2019 were included in this study. Six(30%)out of 20 patients showed fistula. Dysphagia score was improved significantly before and after stenting(3.3± 0.6 vs 1.8±0.9, p<0.01). Complications associated with stenting occurred in 6(30%)cases. Thirteen(65%)patients were able to be discharged from the hospital, but 7(35%)patients including 4 with fistula were outcomes of death in the hospital. Esophageal stenting for malignant esophageal stenosis improved food ingestion and QOL. The prognosis in the case of malignant esophageal stenosis with fistula is extremely poor.

[Application of Sugar to the Edematous Stoma for Obstructive Rectal Cancer-A Case Report].

We herein report a case of application of sugar to the edematous stoma for obstructive rectal cancer. A 70-year-old male patient was diagnosed with rectal cancer, bowel obstruction and multiple lung metastases. Colostomy was performed. Seven days after operation, severe edema and congestion of stoma continued. We started spraying of sugar to stoma, and a few days later, edema and congestion of stoma improved. Before discharge, stoma size markedly reduced. Steady state of stoma and achievement the ability to self-care their stoma is important for introduction of chemotherapy. Application of sugar to reduce edema of rectal prolapse and prolapsed stoma have reported. Although the number of reported cases is still small, effectiveness of sugar to reduce edema of stoma have reported. In our case, application of sugar to the stoma is effective in reduction of edema. Application of sugar might be effective in reduction edema of stoma.

Molecular-scale visualization and surface charge density measurement of Z-DNA in aqueous solution.

The DNA in the left-handed conformation (Z-conformation) was first discovered by A. Rich, who revealed the crystalline structure of a DNA oligomer d(GC)3 by X-ray diffraction method. Later it was also found that DNA molecules change their conformations from typical right-handed form (B-DNA) to the left-handed form (Z-DNA) under specific conditions (B-Z transition). Furthermore, the detailed structures of the interface between B- and Z-DNAs, B-Z junction, was also determined with an atomic resolution. Recently it was found that some proteins have the Z-DNA binding domains, but the biological functions of Z-DNA are not well understood yet. Therefore the investigation of Z-DNA under physiological conditions is highly essential. In this study, we demonstrated the high-resolution real-space imaging of DNA molecules having the Z- and B-form conformations by frequency-modulation atomic force microscopy (FM-AFM), that has made a great progress in recent years, in an aqueous solution. The major and minor grooves of both DNA conformations were clearly visualized. Furthermore, the surface charge density was measured by three-dimensional (3D) force mapping method. We found that Z-form region was less negatively charged than the B-form region.

Author Correction: Atomic-resolution three-dimensional hydration structures on a heterogeneously charged surface.

The original version of the Supplementary Information associated with this Article contained an error in Supplementary Figure 9e,f in which the y-axes were incorrectly labelled from '-40' to '40', rather than the correct '-400' to '400'. The HTML has been updated to include a corrected version of the Supplementary Information.

Immunoactivity of self-assembled antibodies investigated by atomic force microscopy.

Immunoglobulin G (IgG), an antibody, plays a significant role in the immune system, and the functions of IgG molecules have been studied in many research fields such as medicine and engineering. Recently, we found the self-assembly of monoclonal mouse IgG molecules on a mica substrate using atomic force microscopy (AFM); the IgG molecules self-assemble into hexamers and the hexamers form a two-dimensional (2D) crystal. The self-assembly of the IgG molecules is of great interest in terms of the enhancement of the immunoactivity of the antibodies. In this study, we investigated the self-assembly of various IgG molecules on a mica substrate to discuss if the hexamerization of the IgG molecules is a general phenomenon. We also investigated the antigen binding site in the IgG antibody hexamers, and estimated the association rate constant of the self-assembled IgG molecules based on the AFM measurements. The estimated value was lower than that reported in a previous study probably because of the limited mobility of the antigen-binding fragments on the substrate.

Atomic-resolution three-dimensional hydration structures on a heterogeneously charged surface.

Local hydration structures at the solid-liquid interface around boundary edges on heterostructures are key to an atomic-level understanding of various physical, chemical and biological processes. Recently, we succeeded in visualising atomic-scale three-dimensional hydration structures by using ultra-low noise frequency-modulation atomic force microscopy. However, the time-consuming three-dimensional-map measurements on uneven heterogeneous surfaces have not been achieved due to experimental difficulties, to the best of our knowledge. Here, we report the local hydration structures formed on a heterogeneously charged phyllosilicate surface using a recently established fast and nondestructive acquisition protocol. We discover intermediate regions formed at step edges of the charged surface. By combining with molecular dynamics simulations, we reveal that the distinct structural hydrations are hard to observe in these regions, unlike the charged surface regions, possibly due to the depletion of ions at the edges. Our methodology and findings could be crucial for the exploration of further functionalities.

Flexible DNA Path in the MCM Double Hexamer Loaded on DNA.

The formation of the pre-replicative complex (pre-RC) during the G1 phase, which is also called the licensing of DNA replication, is the initial and essential step of faithful DNA replication during the subsequent S phase. It is widely accepted that in the pre-RC, double-stranded DNA passes through the holes of two ring-shaped minichromosome maintenance (MCM) 2-7 hexamers; however, the spatial organization of the DNA and proteins involved in pre-RC formation is unclear. Here we reconstituted the pre-RC from purified DNA and proteins and visualized the complex using atomic force microscopy (AFM). AFM revealed that the MCM double hexamers formed elliptical particles on DNA. Analysis of the angle of binding of DNA to the MCM double hexamer suggests that the DNA does not completely pass through both holes of the MCM hexamers, possibly because the DNA exited from the gap between Mcm2 and Mcm5. A DNA loop fastened by the MCM double hexamer was detected in pre-RC samples reconstituted from purified proteins as well as those purified from yeast cells, suggesting a higher-order architecture of the loaded MCM hexamers and DNA strands.

Thoracoscopic esophagectomy in prone position: advantages of five ports over four ports.

BACKGROUND/AIMS: Thoracoscopic esophagectomy in the prone position (ThE-PP) is usually performed with four ports, which makes the operation almost solo surgery. We now perform ThE-PP with five ports, with the advantage of having the assistant able to provide additional help. The aim of this study was to elucidate the benefits of ThE with five ports over ThE with four ports. METHODOLOGY: We retrospectively reviewed the clinical charts of 47 patients with esophageal cancer who underwent ThE-PP. A total of 14 patients underwent ThE-PP with four ports and 33 with five ports. We compared the number of dissected lymph nodes (LNs)--total; upper left, middle, and lower mediastinum--between the four-port and five-port groups. RESULTS: The number of LNs dissected, including the total, the upper left and middle mediastinum, were not significantly different between the two operations. The number of LNs dissected from the lower mediastinum, however, was significantly higher in the five-port group (median number and interquartile range: 5 and 2-7, respectively) than in the four-port group (0.5 and 0-3, respectively) (P < 0.01). CONCLUSIONS: ThE-PP performed with five ports has an advantage over the same operation done with four ports in terms of lymphadenectomy in the lower mediastinum.

Immunoactive two-dimensional self-assembly of monoclonal antibodies in aqueous solution revealed by atomic force microscopy.

The conformational flexibility of antibodies in solution directly affects their immune function. Namely, the flexible hinge regions of immunoglobulin G (IgG) antibodies are essential in epitope-specific antigen recognition and biological effector function. The antibody structure, which is strongly related to its functions, has been partially revealed by electron microscopy and X-ray crystallography, but only under non-physiological conditions. Here we observed monoclonal IgG antibodies in aqueous solution by high-resolution frequency modulation atomic force microscopy (FM-AFM). We found that monoclonal antibodies self-assemble into hexamers, which form two-dimensional crystals in aqueous solution. Furthermore, by directly observing antibody-antigen interactions using FM-AFM, we revealed that IgG molecules in the crystal retain immunoactivity. As the self-assembled monolayer crystal of antibodies retains immunoactivity at a neutral pH and is functionally stable at a wide range of pH and temperature, the antibody crystal is applicable to new biotechnological platforms for biosensors or bioassays.

Multidetector computed tomography for preoperative identification of left gastric vein location in patients with gastric cancer.

BACKGROUND: The area near the left gastric vein (LGV) is a challenging site at which to perform dissection of the lymph nodes during gastrectomy. Therefore, knowledge of the precise location of the LGV is important. The objective of this study was to examine the usefulness of multidetector computed tomography (MDCT) for the identification of the LGV. METHODS: Eighty-one patients with gastric cancer underwent MDCT, which was performed with contrast media in 76 patients and without contrast media in 5 patients. A 5-mm thin slice of the frontal image was reconstructed. These images were examined preoperatively to detect the location of the LGV. Upon gastrectomy, the LGV was identified and its location compared to that determined by MDCT. RESULTS: The LGV was identified by MDCT in 76 of the 81 patients (93.8%). The LGV was subsequently located during the operation in all 81 patients. The LGV was located dorsal to the common hepatic artery in 40 patients (49.4%), ventral to the common hepatic artery in 18 patients (22.2%), ventral to the splenic artery in 17 patients (21.0%), dorsal to the splenic artery in 2 patients (2.5%) and in other positions in 4 patients (4.9%). In all patients, the location of the LGV detected using MDCT was consistent with that identified during gastrectomy. In the 4 patients with relatively unusual locations of the LGV, these 4 LGV variants were identified preoperatively by MDCT. CONCLUSION: MDCT was useful for identifying the location of the LGV prior to gastrectomy.