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Background: Lung subtraction iodine mapping (LSIM)-CT is a clinically useful technique that can visualize pulmonary mal-perfusion in patients with chronic thromboembolic pulmonary disease (CTEPD). However, little is known about the associations of LSIM images with hemodynamic parameters of patients with CTEPD. This study investigates a parameter of LSIM images associated with mean pulmonary arterial pressure (mPAP) and validates the association between pulmonary vascular resistance, right atrial pressure, cardiac index, and exercise capacity in patients with CTEPD., Methods: This single-center, prospective, observational study involved 30 patients diagnosed with CTEPD using lung perfusion scintigraphy. To examine the correlation of decreased pulmonary perfusion area (DPA) with mPAP, areas with 0-10, 0-15, 0-20, and 0-30 HU in lung subtraction images were adopted in statistical analysis. The DPA to total lung volume ratio (DPA ratio, %) was calculated as the ratio of each DPA volume to the total lung volume. To assess the correlation between DPA ratios of 0-10, 0-15, 0-20, and 0-30 HU and mPAP, Spearman's rank correlation coefficient was used., Results: The DPA ratio of 0-10 HU had the most preferable correlation with mPAP than DPA ratios of 0-15, 0-20, and 0-30 HU ( ρ  = 0.440, P  = 0.015). The DPA ratio of 0-10 HU significantly correlates with pulmonary vascular resistance ( ρ  = 0.445, P  = 0.015). The receiver operating characteristic curve analysis indicated that the best cutoff value of the DPA ratio of 0-10 HU for the prediction of an mPAP of ≥30 mmHg was 8.5% (AUC, 0.773; 95% CI, 0.572-0.974; sensitivity, 83.3%; specificity, 75.0%). Multivariate linear regression analysis, which was adjusted for the main pulmonary arterial to ascending aortic diameter ratio and right ventricular to left ventricular diameter ratio, indicated that the DPA ratio of 0-10 HU was independently and significantly associated with mPAP ( B  = 89.7; 95% CI, 46.3-133.1, P  < 0.001)., Conclusion: The DPA ratio calculated using LSIM-CT is possibly useful for estimating the hemodynamic status in patients with CTEPD., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Yamaguchi, Ehara, Yoshida, Himoto, Izuta, Hayashi, Hayashi, Ogawa, Shibata, Yamazaki, Izumiya and Fukuda.)

Aim: Patients with liver cirrhosis and portosystemic shunt occasionally develop reversed portal flow in the portal venous system. The factors contributing to reversed portal flow in these patients remain unclear. The aim of this study was to identify factors contributing to reversed portal flow in patients with portosystemic shunts based on four-dimensional computed tomography (4DCT), which visualized flow dynamics in the portal venous system., Methods: Data from 34 consecutive patients with portosystemic shunts who had undergone 4DCT before interventional radiology procedures were retrospectively investigated in this study. Uni- and multivariate analyses were performed to identify factors contributing to reversed portal flow., Results: Flow dynamics could be visualized on 4DCT in 32 of the 34 patients. Fifteen patients had forward portal flow; 17 had reversed portal flow. The main portal, splenic, and superior mesenteric veins displayed reversed portal flow in five, 12, and five vessels, respectively. Portosystemic shunt originating from splenic and superior mesenteric veins, worse albumin-bilirubin score, and small main portal vein diameter were significant factors contributing to reversed portal flow in both univariate (p = 0.049, p = 0.027, and p = 0.002) and multivariate (odds ratio [OR] 6.345, p = 0.012; OR 4.279, p = 0.039; and OR 5.516, p = 0.019) analyses., Conclusions: The reversed portal flow was visualized on 4DCT. Portosystemic shunt originating distant to the liver, worse albumin-bilirubin score, and small diameter of the main portal vein were factors contributing to reversed flow in the portal venous system., (© 2022 Japan Society of Hepatology.)

Methods to construct single-cell pairs of heterogeneous cells attract attention because of their potential in cell biological and medical applications for analyzing individual intercellular communications such as immune and nerve synaptic interactions. Photoactivatable substrate surfaces for cell anchoring are promising tools to achieve single-cell pairing. However, conventional surfaces that photoactivate a single type of cell anchoring moiety restrict the combination of cell pair types and their applications. We developed a photoresponsive material comprising a bioorthogonal photoreactive moiety and non-cell adhesive hydrophilic polymer. The material-coated surface allows conjugation with various cell anchoring molecules in response to light at specific timings and consequently achieves light-induced anchoring of a variety of cells at defined regions. Using the platform surface, an array of cancer cell and natural-killer (NK) cell pairs was constructed on a flat substrate surface and the dynamic morphological changes of the cancer cells were monitored by cytotoxic interaction with NK cells at a single-cell level. The photoreactive surface is a useful tool for image-based investigation of the communications between a variety of cell types.

Versatile methods for patterning multiple types of cells with single-cell resolution have become an increasingly important technology for cell analysis, cell-based device construction, and tissue engineering. Here, we present a photoactivatable material based on poly(ethylene glycol) (PEG)-lipids for patterning a variety of cells, regardless of their adhesion abilities. In this study, PEG-lipids bearing dual fatty acid chains were first shown to perfectly suppress cell anchoring on their coated substrate surfaces whereas those with single-chain lipids stably anchored cells through lipid-cell membrane interactions. From this finding, a PEG-lipid with one each of both normal and photocleavable fatty acid chains was synthesized as a material that could convert the chain number from two to one by exposure to light. On the photoconvertible PEG-lipid surface, cell anchoring was activated by light exposure. High-speed atomic force microscopy measurements revealed that this photocaging of the lipid-cell membrane interaction occurs because the hydrophobic dual chains self-assemble into nanoscale structures and cooperatively inhibit the anchoring. Light-induced dissociation of the lipid assembly achieved the light-guided fine patterning of multiple cells through local photoactivation of the anchoring interactions. Using this surface, human natural killer cells and leukemia cells could be positioned to interact one-by-one. The cytotoxic capacity of single immune cells was then monitored via microscopy, showing the proof-of-principle for applications in the high-throughput analysis of the heterogeneity in individual cell-cell communications. Thus, the substrate coated with our photoactivatable material can serve as a versatile platform for the accurate and rapid patterning of multiple-element cells for intercellular communication-based diagnostics.

Objectives: Cases in which the left circumflex coronary artery (LCX) runs close to the mitral annulus are considered high risk for LCX injury during mitral surgery. We investigated the anatomical relationship between the LCX and the mitral annulus using 3-dimensional (3D) computed tomography (CT)., Methods: We constructed 3D-CT images of the LCX and the mitral annulus before surgery in 122 patients with mitral regurgitation (MR). We classified coronary dominance by 3D-CT and MR aetiologies (degenerative, atrial functional MR, ventricular functional MR and Barlow's disease) using echocardiography. We detected the point on the mitral annulus closest to the LCX (X point) and measured the minimum distance from the LCX to the mitral annulus (mCAD). We judged whether atrioventricular disjunction existed using CT. We also investigated the factors affecting mCAD and examined how coronary dominance and MR aetiologies relate to the location of the X point., Results: The median mCAD was 4.2 mm (range 0.9-11.4 mm). Considering coronary dominance and MR aetiologies, mCAD was shorter in patients with left coronary dominance and Barlow's disease. The X point mostly existed on the lateral side of the posterior annulus, but it sometimes existed on the medial side. Multiple regression revealed left dominance and mitral annular disjunction as significant factors affecting mCAD (P = 0.01)., Conclusions: The anatomical relationship between the LCX and the mitral annulus can be recognized using superimposed 3D-CT images. This approach is useful to avoid LCX injury in mitral valve surgery since the X point varies between patients., (© The Author(s) 2021. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery. All rights reserved.)

Multidetector row computed tomography (CT) scanners perform dynamic scanning and have a wide scan range. Time-resolved three-dimensional CT (i.e., 4D CT) has recently enabled visualization of flow in neurovascular vessels. We hypothesized that 4D CT technology would be a useful and non-invasive method for visualizing the flow dynamics of the portal circulation. The aim of this study was to evaluate the technical feasibility of 4D CT for visualizing flow dynamics in the portal circulation using 320-detector-row CT. 4D CT images of 18 consecutive patients with portal circulation including gastrorenal shunt were retrospectively evaluated for their ability to generate flow dynamics of the portal circulation. Flow dynamics could be visualized by 4D CT in 68 of the 72 vessels in the portal vein, splenic vein, superior mesenteric vein, and gastrorenal shunt. Flow direction could not be identified in four vessels, all of them being superior mesenteric veins. Flow direction was recognized on 4D CT in the 68 vessels of the portal circulation. A preliminary validation study revealed that flow direction of all 19 vessels in the portal circulation had concordance between 4D CT and color Doppler ultrasound. 4D CT could visualize flow dynamics of the portal circulation.

We report a photo-cleavable material for tight trapping of nonadherent cells to substrate surfaces. Model immunocytes were selectively trapped in a non-irradiated area as single cells after the projection of a light pattern and withstood high-speed laminar flow, achieving light-guided cell release from the substrates.

Ketamine has been used to treat chronic pain; however, it is still unknown as to what types of chronic pain is ketamine effective against. To identify the effect of administration of subanesthetic-dose ketamine in patients with chronic pain and to clarify the mechanism of the effect, we retrospectively investigated brain functional connectivity using resting-state functional magnetic resonance imaging (rs-fMRI). Patients were divided into responders (Group R: ≥50% improvement on Numerical Rating Scale) and non-responders (Group NR). We compared the differences in terms of brain functional connectivity by seed-to-voxel correlation analysis. Two-sample t-test revealed significant lower connectivity between the medial prefrontal cortex (mPFC) and precuneus in Group R. We also found a significant negative correlation between the improvement rate and functional connectivity strength between the mPFC and precuneus. These findings suggest that subanesthetic-dose ketamine is effective in patients with chronic pain whose brain functional connectivity between the mPFC and precuneus is low. We believe that the current study explored for the first time the correlation between brain functional connectivity and the effect of subanesthetic-dose ketamine for chronic pain and indicated the possibility of use of the predictive marker in pharmacological treatment of chronic pain.

Spatiotemporal control of cell-material interactions contributes to our understanding of cell biology and the development of cell engineering. Here, we first report the reversible and spatio-selective immobilization of nonadherent cells through the use of photoswitchable polymeric materials. The substrate coated with spiropyran-conjugated poly(ethylene glycol) (PEG) lipids, which bind to cell membranes via the lipid moiety only in their merocyanine form, enabled rapid cell immobilization and release in an on-off manner by irradiation with ultraviolet and visible light, respectively. Our work has the potential to improve the performance of cell manipulations on chips and to enable rapid cell arrangement/sorting on various surfaces.

Aim: To investigate the anesthetic management of peroral endoscopic myotomy (POEM) and its associated complications., Methods: This study was a single-center, retrospective, observational study comprising a case series of all patients who underwent POEM in our hospital from April 2015 to November 2016. We collected data regarding patient characteristics, anesthetic methods, surgical factors, and complications using an electronic chart., Results: There were 86 patients who underwent POEM in our hospital during the study period. Preoperatively, patients were maintained on a low residue diet for 48 h prior to the procedure. They were fasted of solids for 24 h before surgery. There was one case of aspiration (1.2%). During POEM, patients were positioned supine with the upper abdomen covered by a clear drape so that pneumoperitoneum could be timeously identified. In three cases, the peak airway pressure exceeded 35 cmH 2 O during volume controlled ventilation with tidal volumes of 6-8 mL/kg and subsequent impairment of ventilation. These cases had been diagnosed with spastic esophageal disorders (SEDs) and the length of the muscular incision on the esophageal side was longer than normal., Conclusion: In the anesthetic management of POEM, it is important to prevent aspiration during induction of anesthesia and to identify and treat complications associated with CO 2 insufflation., Competing Interests: Conflict-of-interest statement: No potential conflicts of interest relevant to this article were reported.

In a search for compounds interacting with ergosterol resin, a new compound named dipyrimicin B was isolated from a rare actinomycete strain, Amycolatopsis sp. K16-0194. In addition, another analog, dipyrimicin A, which does not interact with the resin, was also discovered. The structures of the two dipyrimicins were established by comprehensive 1D and 2D NMR and MS analyses and found to contain a unique core structure, a 2,2'-bipyridine skeleton. Dipyrimicin A showed strong antimicrobial and cytotoxic activity, whereas dipyrimicin B displayed distinctly poor antimicrobial and cytotoxic activities.

ᅟ: We report a case of unexpected ventilatory impairment that occurred during per-oral endoscopic myotomy (POEM) under general anesthesia. A 73-year-old woman underwent POEM for Jackhammer esophagus. The patient developed hypercarbia, pneumoperitoneum, and severe subcutaneous emphysema during the operation. Although she was treated with abdominal paracentesis, it became difficult to ventilate her lungs a few minutes later. We recommended the surgeons to interrupt the procedure and proposed repeating the abdominal paracentesis. Simultaneously, we switched to manual ventilation and waited for the subcutaneous emphysema to subside. Thereafter, her respiratory status gradually improved and the surgeons were able to continue the operation. We considered that the main reason for our patient's severe ventilatory impairment was that the length of surgical dissection was longer than usual.

Molecular networks on the cytoplasmic faces of cellular plasma membranes are critical research topics in biological sciences and medicinal chemistry. However, the selective permeability of the cell membrane restricts the researchers from accessing to the intact intracellular factors on the membrane from the outside. Here, a microfluidic method to prepare cell membrane sheets was developed as a promising tool for direct examination of the cytoplasmic faces of cell membranes. Mammalian cells immobilized on a poly(ethylene glycol)-lipid coated substrate were rapidly and efficiently fractured, with the sheer stress of laminar flow in microchannels, resulting in isolation of the bottom cell membrane sheets with exposed intact cytoplasmic faces. On these faces of the cell membrane sheets, both ligand-induced phosphorylation of receptor tyrosine kinases and selective enzymatic modification of a G-protein coupling receptor were directly observed. Thus, the present cell membrane sheet should serve as a unique platform for studies providing new insights into juxta-membrane molecular networks and drug discovery.

A chemically-activatable alkynyl steroid analogue probe has been synthesized for visualizing the lipid raft membrane domains by Raman microscopy. The Raman probe, in which ring A of its steroid backbone is replaced with an alkynyl group, was designed to enable activation of the alkyne signal through the Eschenmoser-Tanabe fragmentation reaction of the oxidized cholesterol precursor in lipid bilayer membranes. The alkynyl steroid analogue was observed to form liquid-ordered raft-like domains on a model giant-liposome system in a similar manner as cholesterol, and the large alkyne signal of the accumulated probe at 2120 cm -1 was mapped on the microdomains with a Raman microscope. The alkyne moiety of the probe was confirmed to be converted from the α,β-epoxy ketone group of its precursor by reaction with p-toluensulfonyl hydrazine under a mild condition. Through the reaction, the alkyne signal of the probe was activated on the lipid bilayer membrane of liposomes. Furthermore, the signal activation of the probe was also detected on living cells by stimulated Raman scattering microscopy. The ring-A-opened alkyne steroid analogue, thus, provides a first chemically-activatable Raman probe as a promising tool for potentially unravelling the intracellular formation and trafficking of cholesterol-rich microdomains.

Stimuli-responsive "activatable" reactive tags are applicable to selective labeling of biomolecules in a defined area or environment in living systems, yielding new insights into cellular processes through molecular imaging and fishing. Here, we developed a chemically activatable alkyne tag that can be incorporated into biological molecules and labeled with azide-tagged functional molecules through the alkyne-azide cycloaddition "click" reaction after chemical activation. Formation of the alkyne tag from the precursor moiety was confirmed to proceed in physiological aqueous media and was particularly enhanced under mildly acidic pH. The tag was successfully applied to low-pH sensitive labeling of a cholesterol analogue with azide-tagged biotin on living mammalian cells. Our results provided proof of principle that the present activatable alkyne tag can be used as a tool to selectively analyze molecules of interest in low-pH regions in living systems.

Background: There is few study to examine the vital sign changes and the requirement of analgesics after discontinuation of dexmedetomidine in postopera- tive patients., Methods: This is a retrospective observational study conducted in 74 patients after esophageal cancer sur- gery. We recorded vital signs including blood pressure, heart rate and respiratory rate one hour before discon- tinuation of dexmedetomidine, and at 1, 2, 4, 6 hours after its discontinuation. We also recorded the use of opioid and analgesic within 6 hours after discontinua- tion., Results: Mean blood pressure, pulse rate, and respi- ratory rate significantly increased after DEX discon- tinuation. Compared with the data before discontinua- tion, the mean blood pressure increased by 13.3 mmHg, heart rate increased by 7.5 beats - min- and respira- tory rate increased by 3.0 times - min-' in average at 6 hours after discontinuation. There were 28 patients (38%) who required the additional analgesics within 6 hours after discontinuation., Conclusions: After discontinuation of dexmedetomi- dine, significant changes of vital signs, especially in mean blood pressure, were observed in post-esopha- gectomy patients. About 38% of them required addi- tional analgesics within 6 hours after dexmedetomidine discontinuation.

We report a case of delayed respiratory depression due to accidental subcutaneous opioid infusion. A healthy 33-year-old woman underwent orthopedic surgery under general anesthesia. Before the end of the operation, it was noticed that a part of the opioid infusion had been administered subcutaneously. About 15 min after tracheal extubation, the patient developed respiratory depression and loss of consciousness. The patient recovered with the use of jaw lift together with bag-valve-mask ventilation. We believe that accidental subcutaneous opioid accumulation may have caused the respiratory depression.

We report on the methods and experiences of the dual-phase cone beam computed tomography during hepatic arteriography (CBCTHA) to apply the 3D-DSA. A total of 32 ml contrast medium (150 mgI/ml) was injected at the rate of 2.0 ml/s for 16 s. The early phase scan was initiated 10 s after the start of contrast media injection. The delayed phase scan was started 40 s after that (24 s after the end of CM injection). When using the dual phase CBCTHA, it was able to obtain the classical hepatocellular carcinoma (HCC) images same as computed tomography during hepatic arteriography (CTHA). In the early phase, the tumor can be highly enhanced against the liver parenchyma. In delayed phase, corona enhancement was clearly appeared at the liver parenchyma. Of 58 cases of acquisitions, we experienced six cases with miss breath holding and 14 cases with over the field of view (FOV) due to hepatomegaly. We evaluated the tumor contrast in 18 cases because the other 40 cases were not applied to our criteria. The pixel values of ROIs on the tumor, coronal enhancement, and liver parenchyma were measured, respectively. Then, we calculated tumor-parenchyma contrast (T-P contrast), corona-tumor contrast (C-T contrast), and corona-parenchyma contrast (C-P contrast). The T-P contrast was 358±112, the C-T contrast was 132±51, and the C-P contrast was 168±66. The contrast was clearly visualized among them. The dual-phase CBCTHA that applies the 3D-DSA is a simple and useful technique for hepatocellular carcinoma treatment.

The many known eukaryotic DNA polymerases are classified into four families; A, B, X, and Y. Among them, DNA polymerase η, a Y family polymerase, is a low fidelity enzyme that contributes to translesional synthesis and somatic hypermutation. Although a high mutation frequency is observed in immunoglobulin genes, translesional synthesis occurs with a high accuracy. We determined whether the misincorporation rate of DNA polymerase η varies with ambient conditions. It has been reported that DNA polymerase η is unable to exclude water molecules from the active site. This finding suggests that some ions affect hydrogen bond formation at the active site. We focused on the effect of pH and evaluated the misincorporation rate of deoxyguanosine triphosphate (dGTP) opposite template T by DNA polymerase η at various pH levels with a synthetic template-primer. The misincorporation rate of dGTP by DNA polymerase η drastically increased at pH 8.0-9.0 compared with that at pH 6.5-7.5. Kinetic analysis revealed that the Km value for dGTP on the misincorporation opposite template T was markedly affected by pH. However, this drastic change was not seen with the low fidelity DNA polymerase α.

The purpose of this study was to understand the scatter radiation distribution during C-arm CT examination in the interventional radiography (IVR) room to show the escaped area and the radiation protective method. The C-arm rotates 200° in 5 s. The tube voltage was 90 kV, and the entrance dose to the detector was 0.36 μGy/frame during C-arm CT examination. The scattered doses were measured each 50 cm from the isocenter like a grid pattern. The heights of the measurement were 50, 100, and 150 cm from the floor. The maximum scattered doses were 38.23±0.60 μGy at 50 cm, 43.86±0.20 μGy at 100 cm, and 25.78±0.37 μGy at 150 cm. The scatter radiation distribution at 100 cm was the highest scattered dose. The operator should protect their reproductive gland, thyroid, and lens. The scattered dose was low behind the C-arm body and the bed, so they will be able to become the escaped area for staff.

The effect of T-5224, a selective inhibitor of c-Fos/activator protein (AP)-1, on lipopolysaccharide (LPS) induced liver injury was examined in mice. Administration of LPS (10 mg kg(-1), i.p.) markedly increased serum levels of tumor necrosis factor-alpha (TNFα), high mobility group box 1 (HMGB1), alanine aminotransferase/aspartate aminotransferase (ALT/AST), liver tissue levels of macrophage-inflammatory protein-1 alpha (MIP-1α) and monocyte chemoattractant protein-1 (MCP-1), as well as hepatic necrosis and inflammation, leading to 67 % lethality. Administration of T-5224 (300 mg kg(-1), p.o.) after intraperitoneal injection of LPS imparted appreciable protection against acute elevations in serum levels of TNFα, HMGB1, ALT/AST as well as in liver tissue levels of MIP-1α and MCP-1, and reduced the lethality (27 %). These data indicate that T-5224 ameliorates liver injury and improves survival through decreasing production of proinflammatory cytokines and chemokines in endotoxemic mice.

A 17-year-old man was scheduled for ventriculo-peritoneal shunt under general anesthesia. Anesthesia was induced by propofol, rocuronium and pentazocine, and maintained with sevoflurane and nitrous oxide. The operation was finished in 1 hour and 4 minutes without trouble. After the operation, sugammadex was administered. After about 2 minutes, the redness and the wheal appeared in the body trunk, and SpO2 dropped to The appearance of rapid symptom immediately after administration of sugammadex indicated that anaphylactic reaction had occurred. We administered adrenaline. After the administration of adrenaline several times, cutaneous findings became improved and SpO2 was stabilized. We started continuous infusion of adrenaline, and the patient was moved to the intensive care unit (ICU). Twelve hours after entering the ICU, the patient was extubated. He showed no troubles thereafter, and left hospital. After introduction of sugammadex to clinical use, 7 cases of anaphylactic reaction to sugammadex were reported. We must know that anaphylactic reaction can be induced by sugammadex.

Translesion DNA synthesis (TLS) involves PCNA mono-ubiquitination and TLS DNA polymerases (pols). Recent evidence has shown that the mono-ubiquitination is induced not only by DNA damage but also by other factors that induce stalling of the DNA replication fork. We studied the effect of spontaneous DNA replication errors on PCNA mono-ubiquitination and TLS induction. In the pol1L868F strain, which expressed an error-prone pol alpha, PCNA was spontaneously mono-ubiquitinated. Pol alpha L868F had a rate-limiting step at the extension from mismatched primer termini. Electron microscopic observation showed the accumulation of a single-stranded region at the DNA replication fork in yeast cells. For pol alpha errors, pol zeta participated in a generation of +1 frameshifts. Furthermore, in the pol1L868F strain, UV-induced mutations were lower than in the wild-type and a pol delta mutant strain (pol3-5DV), and deletion of the RAD30 gene (pol eta) suppressed this defect. These data suggest that nucleotide misincorporation by pol alpha induces exposure of single-stranded DNA, PCNA mono-ubiquitination and activates TLS pols.

DNA polymerase eta is recently found as a responsible gene product of xeroderma pigmentosum variant. Differently from other eukaryotic DNA polymerases, such as alpha, beta or gamma, eta polymerase is categorized in Y family. Specific inhibitors for DNA polymerases are useful tools to study the exact role of enzyme in the living cells, however, the inhibitor for DNA polymerase eta has not been developed. We examined the inhibitory effects of several sugar-modified nucleotide analogs on DNA polymerase eta. The arabinonucleotides (araCTP), dideoxynucleotides (ddTTP) and 3'-modified nucleotides (3'-dCTP and 3'-azidothymidine tri-phosphate) did not show any inhibitory effect on DNA polymerase eta. On the other hand, the oxetanocin derivatives those have the oxetane ring as a sugar moiety, OXT-GTP and OXT-ATP, strongly inhibited this polymerase. These results suggest that DNA polymerase eta has a unique recognition mode against the sugar moiety of nucleotide compared with other eukaryotic nucleotide polymerases.

Replication protein A (RAP) is a eukaryotic single-stranded DNA binding protein involved in DNA replication, repair, and recombination. Recent studies indicate that RPA preferentially binds the damaged sites rather than the undamaged sites. Therefore, RPA is thought to be a member ofrepair factories or a sensor of lesion on DNA. To obtain further information of behavior of RPA against the oxidized lesion, we studied the binding affinity of RPA for the single-stranded DNA containing 5-formyluracil, a major lesion of thymine base yielded by the oxidation, using several synthetic oligonucleotides. The affinity of RPA for oligonucleotides was determined by gel shift assay. Results suggest that the surrounding sequence of 5-formyluracil may affect the affinity for RPA, and that the 5-formyluracil on the purine stretch but not the pyrimidine stretch increases the affinity for RPA. Results of affinity labeling experiment of RPA with the oligonucleotides containing 5-formyluracil indicate that RPA1 subunit may directly recognize and bind to the 5-formyluracil on the single-stranded DNA.

To clarify the molecular mechanism of mismatch formation by DNA polymerase eta, the pH-dependency of misincorporation of dNTP was studied with the synthetic template-primer. Incorporation of dNTP formed Watson-Crick type base pair, such as the incorporation of dATP opposite template T, was slightly affected by pH between 6.5 to 9.0. On the other hand, the misincorporation rate of dGTP opposite template T by DNA polymerase eta was drastically increased according to the increasing pH. Kinetical analysis revealed that this change might be due to the change of Km value for dGTP rather than that of Vmax value. This suggests that the affinity of dGTP on DNA polymerase eta during the mismatch formation with template T should be affected by pH.

DNA polymerase is one of the most important target molecules of antitumor agents, especially for antimetabolite nucleosides that include 1-beta-D-arabinofuranosylcytosine (araC) and 2'-deoxy-2',2'-difluorocytidine (gemcitabine). There are several subtypes of mammalian DNA polymerases and their localization and function have been clarified. DNA polymerase alpha, delta and epsilon have been implicated to be responsible for DNA replication, whereas DNA polymerase beta, delta and epsilon have been suggested to work in DNA repair. DNA polymerase gamma is encoded in the nucleus but localizes in the mitochondria, and is responsible for the mitochondrial DNA replication. Recently, several antiviral nucleoside analogs were reported to inhibit DNA polymerase gamma after intracellular phosphorylation and cause severe chronic toxicity. 1-(2-Deoxy-2-fluoro-4-thio-beta-D-arabinofuranosyl)cytosine (4'-thio-FAC), an antimetabolite similar to araC and gemcitabine, is recently shown by us to be a very promising agent because of its potent antitumor activity by oral administration to mice. We tested for the inhibitory activities of the triphosphates of 4'-thio-FAC and gemcitabine against DNA polymerase alpha, beta and gamma. The triphosphates of 4'-thio-FAC (4'-thio-FACTP) exhibited the potent inhibitory action against DNA polymerase alpha, whereas it showed moderate inhibition against DNA polymerase beta and little inhibition against DNA polymerase gamma. The triphosphate of gemcitabine (dFdCTP) did not show potent inhibition against these three DNA polymerases. Thus, the effect on ribonucleotide reductase was suggested to be more responsible for the antitumor action of gemcitabine. The differences in the mechanisms of action against DNA polymerases between these drugs and other nucleosides were also discussed.

Pulmonary thromboembolism (PTE) is a crucial complication characterized by rapid onset and a high mortality rate. There are few reports concerning perioperative incidence of PTE and deep vein thrombosis (DVT) among Japanese population. All surgical patients excluding young patients for minor surgery were equipped with calf- or thigh-length intermittent pneumatic compression (IPC) device as a DVT preventive method during the period of immobility. Incidence and clinical characteristics of perioperative PTE and DVT under the use of IPC device were analyzed at Kobe University Hospital. One patient developed DVT and four patients developed PTE among the 6500 surgical cases. All patients with DVT or PTE were 66 years or older, and all patients were diagnosed by computer tomography, venography and pulmonary angiography between 7 to 14 days after each operation. One patient was surgically treated by embolectomy and removal of intraarterial thrombus, and others were treated with thrombolysis, anticoagulants and placement of vena cava filters. For prevention of DVT and PTE, other preventive methods such as anticoagulants and prolonged application of IPC should be necessary.