An artificial nucleoside that simultaneously detects and combats drug resistance to doxorubicin.

OBJECTIVES: Doxorubicin is a DNA-damaging agent used to treat hematological cancers. Unfortunately, drug resistance can occur by defective DNA repair activity coupled with the ability of DNA polymerases to misreplicate unrepaired DNA lesions. This study demonstrates that the efficacy of doxorubicin can be improved by using an artificial nucleoside to efficiently and selectively inhibit this activity. METHODS: In vitro studies using acute lymphoblastic leukemia cell lines define the mechanism of cell death caused by combining an artificial nucleoside with doxorubicin. RESULTS: Flow cytometry experiments demonstrate that combining an artificial nucleoside with doxorubicin potentiates the cell killing effects of the drug by increasing apoptosis. The potentiation effect correlates with expression of TdT, a specialized DNA polymerase overexpressed in acute lymphoblastic leukemia. Cell cycle experiments demonstrate that this combination blocks cells at S-phase prior to inducing apoptosis. Finally, the unique chemical composition of the nucleoside analog was used to visualize the replication of damaged DNA in TdT-positive cells. This represents a potential diagnostic tool to easily identify doxorubicin-resistant cancer cells. CONCLUSION: Studies demonstrate that a novel artificial nucleoside improves the therapeutic efficacy of doxorubicin, thereby reducing the risk of potential side effects caused by the DNA-damaging agent.

The use of modified and non-natural nucleotides provide unique insights into pro-mutagenic replication catalyzed by polymerase eta.

This report evaluates the pro-mutagenic behavior of 8-oxo-guanine (8-oxo-G) by quantifying the ability of high-fidelity and specialized DNA polymerases to incorporate natural and modified nucleotides opposite this lesion. Although high-fidelity DNA polymerases such as pol δ and the bacteriophage T4 DNA polymerase replicating 8-oxo-G in an error-prone manner, they display remarkably low efficiencies for TLS compared to normal DNA synthesis. In contrast, pol η shows a combination of high efficiency and low fidelity when replicating 8-oxo-G. These combined properties are consistent with a pro-mutagenic role for pol η when replicating this DNA lesion. Studies using modified nucleotide analogs show that pol η relies heavily on hydrogen-bonding interactions during translesion DNA synthesis. However, nucleobase modifications such as alkylation to the N2 position of guanine significantly increase error-prone synthesis catalyzed by pol η when replicating 8-oxo-G. Molecular modeling studies demonstrate the existence of a hydrophobic pocket in pol η that participates in the increased utilization of certain hydrophobic nucleotides. A model is proposed for enhanced pro-mutagenic replication catalyzed by pol η that couples efficient incorporation of damaged nucleotides opposite oxidized DNA lesions created by reactive oxygen species. The biological implications of this model toward increasing mutagenic events in lung cancer are discussed.

A Genetic Selection for dinB Mutants Reveals an Interaction between DNA Polymerase IV and the Replicative Polymerase That Is Required for Translesion Synthesis.

Translesion DNA synthesis (TLS) by specialized DNA polymerases (Pols) is a conserved mechanism for tolerating replication blocking DNA lesions. The actions of TLS Pols are managed in part by ring-shaped sliding clamp proteins. In addition to catalyzing TLS, altered expression of TLS Pols impedes cellular growth. The goal of this study was to define the relationship between the physiological function of Escherichia coli Pol IV in TLS and its ability to impede growth when overproduced. To this end, 13 novel Pol IV mutants were identified that failed to impede growth. Subsequent analysis of these mutants suggest that overproduced levels of Pol IV inhibit E. coli growth by gaining inappropriate access to the replication fork via a Pol III-Pol IV switch that is mechanistically similar to that used under physiological conditions to coordinate Pol IV-catalyzed TLS with Pol III-catalyzed replication. Detailed analysis of one mutant, Pol IV-T120P, and two previously described Pol IV mutants impaired for interaction with either the rim (Pol IVR) or the cleft (Pol IVC) of the ß sliding clamp revealed novel insights into the mechanism of the Pol III-Pol IV switch. Specifically, Pol IV-T120P retained complete catalytic activity in vitro but, like Pol IVR and Pol IVC, failed to support Pol IV TLS function in vivo. Notably, the T120P mutation abrogated a biochemical interaction of Pol IV with Pol III that was required for Pol III-Pol IV switching. Taken together, these results support a model in which Pol III-Pol IV switching involves interaction of Pol IV with Pol III, as well as the ß clamp rim and cleft. Moreover, they provide strong support for the view that Pol III-Pol IV switching represents a vitally important mechanism for regulating TLS in vivo by managing access of Pol IV to the DNA.

Visualizing nucleic acid metabolism using non-natural nucleosides and nucleotide analogs.

Nucleosides and their corresponding mono-, di-, and triphosphates play important roles in maintaining cellular homeostasis. In addition, perturbations in this homeostasis can result in dysfunctional cellular processes that cause pathological conditions such as cancer and autoimmune diseases. This review article discusses contemporary research areas applying nucleoside analogs to probe the mechanistic details underlying the complexities of nucleoside metabolism at the molecular and cellular levels. The first area describes classic and contemporary approaches used to quantify the activity of nucleoside transporters, an important class of membrane proteins that mediate the influx and efflux of nucleosides and nucleobases. A focal point of this section is describing how biophotonic nucleosides are replacing conventional assays employing radiolabeled substrates to study the mechanism of these proteins. The second section describes approaches to understand the utilization of nucleoside triphosphates by cellular DNA polymerases during DNA synthesis. Emphasis here is placed on describing how novel nucleoside analogs such as 5-ethynyl-2'-deoxyuridine are being used to quantify DNA synthesis during normal replication as well as during the replication of damaged DNA. In both sections, seminal research articles relevant to these areas are described to highlight how these novel probes are improving our understanding of these biological processes. This article is part of a Special Issue entitled: Physiological Enzymology and Protein Functions.

A metal-containing nucleoside that possesses both therapeutic and diagnostic activity against cancer.

Nucleoside transport is an essential process that helps maintain the hyperproliferative state of most cancer cells. As such, it represents an important target for developing diagnostic and therapeutic agents that can effectively detect and treat cancer, respectively. This report describes the development of a metal-containing nucleoside designated Ir(III)-PPY nucleoside that displays both therapeutic and diagnostic properties against the human epidermal carcinoma cell line KB3-1. The cytotoxic effects of Ir(III)-PPY nucleoside are both time- and dose-dependent. Flow cytometry analyses validate that the nucleoside analog causes apoptosis by blocking cell cycle progression at G2/M. Fluorescent microscopy studies show rapid accumulation in the cytoplasm within 4 h. However, more significant accumulation is observed in the nucleus and mitochondria after 24 h. This localization is consistent with the ability of the metal-containing nucleoside to influence cell cycle progression at G2/M. Mitochondrial depletion is also observed after longer incubations (Δt ∼48 h), and this effect may produce additional cytotoxic effects. siRNA knockdown experiments demonstrate that the nucleoside transporter, hENT1, plays a key role in the cellular entry of Ir(III)-PPY nucleoside. Collectively, these data provide evidence for the development of a metal-containing nucleoside that functions as a combined therapeutic and diagnostic agent against cancer.

Insights into antiamyloidogenic properties of the green tea extract (-)-epigallocatechin-3-gallate toward metal-associated amyloid-ß species.

Despite the significance of Alzheimer's disease, the link between metal-associated amyloid-ß (metal-Aß) and disease etiology remains unclear. To elucidate this relationship, chemical tools capable of specifically targeting and modulating metal-Aß species are necessary, along with a fundamental understanding of their mechanism at the molecular level. Herein, we investigated and compared the interactions and reactivities of the green tea extract, (-)-epigallocatechin-3-gallate [(2R,3R)-5,7-dihydroxy-2-(3,4,5-trihydroxyphenyl)-3,4-dihydro-2H-1-benzopyran-3-yl 3,4,5-trihydroxybenzoate; EGCG], with metal [Cu(II) and Zn(II)]-Aß and metal-free Aß species. We found that EGCG interacted with metal-Aß species and formed small, unstructured Aß aggregates more noticeably than in metal-free conditions in vitro. In addition, upon incubation with EGCG, the toxicity presented by metal-free Aß and metal-Aß was mitigated in living cells. To understand this reactivity at the molecular level, structural insights were obtained by ion mobility-mass spectrometry (IM-MS), 2D NMR spectroscopy, and computational methods. These studies indicated that (i) EGCG was bound to Aß monomers and dimers, generating more compact peptide conformations than those from EGCG-untreated Aß species; and (ii) ternary EGCG-metal-Aß complexes were produced. Thus, we demonstrate the distinct antiamyloidogenic reactivity of EGCG toward metal-Aß species with a structure-based mechanism.

Identification and Characterization of Baicalin as a Phosphodiesterase 4 Inhibitor.

Asthma is a chronic inflammatory disease of lung airways, and pharmacological inhibitors of cyclic adenosine monophosphate-specific phosphodiesterase 4 (PDE4) have been considered as therapeutics for the treatment of asthma. However, development of PDE4 inhibitors in clinical trials has been hampered because of the severe side effects of non-selective PDE4 inhibitors. Here, screening of a plant extract library in conjunction with dereplication technology led to identification of baicalin as a new type of PDE4-selective inhibitor. We demonstrated that while rolipram inhibited the enzyme activity of a range of PDE4 subtypes in in vitro enzyme assays, baicalin selectively inhibited the enzyme activity of PDE4A and 4B. In addition, baicalin suppressed lipopolysaccharide-induced TNF-α expression in macrophage where PDE4B plays a key role in lipopolysaccharide-induced signaling. Furthermore, baicalin treatment in an animal model of allergic asthma reduced inflammatory cell infiltration and TNF-α levels in bronchoalveolar lavage fluids, indicating that the antiinflammatory effects of baicalin in vivo are attributable, in part, to its ability to inhibit PDE4.

Nanoporous Carbon Materials as Solid Contacts for Microneedle Ion-Selective Sensors.

Continuous sensing of biomarkers, such as potassium ions or pH, in wearable patches requires miniaturization of ion-selective sensor electrodes. Such miniaturization can be achieved by using nanostructured carbon materials as solid contacts in microneedle-based ion-selective and reference electrodes. Here we compare three carbon materials as solid contacts: colloid-imprinted mesoporous (CIM) carbon microparticles with ∼24-28 nm mesopores, mesoporous carbon nanospheres with 3-9 nm mesopores, and Super P carbon black nanoparticles without internal porosity but with textural mesoporosity in particle aggregates. We compare the effects of carbon architecture and composition on specific capacitance of the material, on the ability to incorporate ion-selective membrane components in the pores, and on sensor performance. Functioning K+ and H+ ion-selective electrodes and reference electrodes were obtained with gold-coated stainless-steel microneedles using all three types of carbon. The sensors gave near-Nernstian responses in clinically relevant concentration ranges, were free of potentially detrimental water layers, and showed no response to O2. They all exhibited sufficiently low long-term potential drift values to permit calibration-free, continuous operation for close to 1 day. In spite of the different specific capacitances and pore architecture of the three types of carbon, no significant difference in potential stability for K+ ion sensing was observed between electrodes that used each material. In the observed drift values, factors other than the carbon solid contact are likely to play a role, too. However, for pH sensing, electrodes with CIM as a carbon solid contact, which had the highest specific capacitance and best access to the pores, exhibited better long-term stability than electrodes with the other carbon materials.

Cyclometalated iridium(III) complexes with deoxyribose substituents.

Fundamental study of enzymatic nucleoside transport suffers for lack of optical probes that can be tracked noninvasively. Nucleoside transporters are integral membrane glycoproteins that mediate the salvage of nucleosides and their passage across cell membranes. The substrate recognition site is the deoxyribose sugar, often with little distinction among nucleobases. Reported here are nucleoside analogues in which emissive, cyclometalated iridium(III) complexes are "clicked" to C-1 of deoxyribose in place of canonical nucleobases. The resulting complexes show visible luminescence at room temperature and 77 K with microsecond-length triplet lifetimes. A representative complex is crystallographically characterized. Transport and luminescence are demonstrated in cultured human carcinoma (KB3-1) cells.

Design of small molecules that target metal-A{beta} species and regulate metal-induced A{beta} aggregation and neurotoxicity.

The accumulation of metal ions and amyloid-ß (Aß) aggregates found in the brain of patients with Alzheimer's disease (AD) has been suggested to be involved in AD pathogenesis. To investigate metal-Aß-associated pathways in AD, development of chemical tools to target metal-Aß species is desired. Only a few efforts, however, have been reported. Here, we report bifunctional small molecules, N-(pyridin-2-ylmethyl)aniline (L2-a) and N(1),N(1)-dimethyl-N(4)-(pyridin-2-ylmethyl)benzene-1,4-diamine (L2-b) that can interact with both metal ions and Aß species, as determined by spectroscopic methods including high-resolution NMR spectroscopy. Using the bifunctional compound L2-b, metal-induced Aß aggregation and neurotoxicity were modulated in vitro as well as in human neuroblastoma cells. Furthermore, treatment of human AD brain tissue homogenates containing metal ions and Aß species with L2-b showed disassembly of Aß aggregates. Therefore, our studies presented herein demonstrate the value of bifunctional compounds as chemical tools for investigating metal-Aß-associated events and their mechanisms in the development and pathogenesis of AD and as potential therapeutics.

Left anterior descending artery dissection with retrograde aortic dissection during percutaneous coronary intervention: a case report.

Retrograde catheter-induced coronary artery dissection during percutaneous coronary intervention is an exceedingly rare occurrence, and the likelihood of it extending into the aorta is even more uncommon. Typically, surgical treatment involves aortic root replacement combined with coronary artery bypass grafting. However, in this particular case, a meticulous approach was employed. By carefully guiding wires into the true lumens and placing stents in the proximal left main and left anterior descending arteries, the immediate complications were averted by obstructing the retrograde flow in the false lumen. Subsequently, an off-pump coronary artery bypass was performed using the left internal mammary artery to the left anterior descending artery, without the need to manipulate the aorta. This approach resulted in a short operation time and the absence of any other complications.

Intractable chylous leak after radical esophagectomy treated with radiotherapy.

Postoperative chylous leak after esophagectomy is a rare but potentially life-threatening complication that results in hypovolemia, electrolyte imbalance, malnutrition, and immunologic deficiency. However, the management of postoperative chylous leak remains controversial. Following a diagnosis of esophageal cancer, a 64-year-old man was treated by video-assisted thoracoscopic esophagectomy, laparoscopic gastric tube formation, prophylactically thoracic duct ligation, and reconstruction with esophagogastrostomy at the neck level. Massive postoperative drainage from the thorax and abdomen did not initially meet the diagnostic criteria for chylothorax, which was ultimately diagnosed 3 weeks after the operation. Despite various treatments including total parenteral nutrition, octreotide and midodrine, reoperation (thoracic duct ligation and mechanical pleurodesis), and thoracic duct embolization, the chylous leak persisted. Finally, low-dose radiation therapy was administered with a daily dose of 2 Gy and completed at a total dose of 14 Gy. After this, the amount of pleural effusion gradually decreased over 2 weeks, and the last drainage tube was removed. The patient was alive and well at 60 months postoperatively. Herein, we describe a patient with intractable chylous leak after esophagectomy, which persisted despite conservative treatment, thoracic duct ligation, and embolization, but was finally successfully treated with radiotherapy.

Large mesenchymal cystic and chondroid pulmonary hamartoma mimicking lung cancer: Case report.

Pulmonary hamartoma is the most commonly resected benign neoplasm of lung. The mesenchymal cystic subtype is a rare and often bilaterally occurring variant composed of multiple cysts and nodules. Herein, we present an asymptomatic 70-year-old woman with a large and mostly cystic growth of right hilar region. Computed tomography of the chest and fluorodeoxyglucose positron emission tomography/computed tomography imaging traced its origins to right middle lobe. Overall features suggested primary lung cancer or perhaps other cystic lung disease.Because transbronchial lung biopsy failed to establish a histologic diagnosis, right middle lobectomy was undertaken by video-assisted thoracoscopic surgery. The gross surgical specimen harbored a single and sizeable (8.0 × 4.0 cm) cystic lesion containing multiple yellow-white nodules. A diagnosis of mesenchymal cystic and chondroid hamartoma was ultimately rendered. This particular case is noteworthy, given the initial clinical resemblance to primary lung cancer.

A Comparative Study on the Taste Characteristics of Satellite Cell Cultured Meat Derived from Chicken and Cattle Muscles.

This study investigated the amino acid and nucleotide-related compound composition and taste characteristics of cultured muscle tissue (CMT) obtained by culturing satellite cells isolated from chicken and cattle and compared them to those of traditional meat (TM). The content of all amino acids except valine and tyrosine was significantly different between CMT and TM (p<0.05). The amount of glutamic acid was not significantly different between CMT and TM in cattle, but the glutamic acid in chicken CMT was lower than that of TM (p<0.05). Among the nucleotide-related compounds, only the content of inosine-5'-monophosphate (IMP) was significant, and the amount of IMP in CMT derived from chicken and cattle was significantly lower than that of TM (p<0.05). There were significant differences in the taste characteristics assessed by an electronic tongue system, and the umami, bitterness, and sourness values of CMT were significantly lower than those of TM from both chicken and cattle (p<0.05). The results of the present study suggest that it is necessary to develop a satellite cell culture method that could increase the umami and bitterness intensity of CMT and adjust the composition of the growth medium to produce cultured meat with a taste similar to that of TM.

Development of bifunctional stilbene derivatives for targeting and modulating metal-amyloid-ß species.

Amyloid-ß (Aß) peptides and their metal-associated aggregated states have been implicated in the pathogenesis of Alzheimer's disease (AD). Although the etiology of AD remains uncertain, understanding the role of metal-Aß species could provide insights into the onset and development of the disease. To unravel this, bifunctional small molecules that can specifically target and modulate metal-Aß species have been developed, which could serve as suitable chemical tools for investigating metal-Aß-associated events in AD. Through a rational structure-based design principle involving the incorporation of a metal binding site into the structure of an Aß interacting molecule, we devised stilbene derivatives (L1-a and L1-b) and demonstrated their reactivity toward metal-Aß species. In particular, the dual functions of compounds with different structural features (e.g., with or without a dimethylamino group) were explored by UV-vis, X-ray crystallography, high-resolution 2D NMR, and docking studies. Enhanced bifunctionality of compounds provided greater effects on metal-induced Aß aggregation and neurotoxicity in vitro and in living cells. Mechanistic investigations of the reaction of L1-a and L1-b with Zn(2+)-Aß species by UV-vis and 2D NMR suggest that metal chelation with ligand and/or metal-ligand interaction with the Aß peptide may be driving factors for the observed modulation of metal-Aß aggregation pathways. Overall, the studies presented herein demonstrate the importance of a structure-interaction-reactivity relationship for designing small molecules to target metal-Aß species allowing for the modulation of metal-induced Aß reactivity and neurotoxicity.

The Korean version of the Virtual Patient Learning System Evaluation Tool: Assessment of reliability and validity.

BACKGROUND: Due to the recent spread of coronavirus disease 2019, Korean nursing colleges are increasingly using virtual patient simulation to make up for a lack of available clinical practice in medical institutions. Therefore, an instrument is required to evaluate the effects of the virtual patient learning system in South Korea. OBJECTIVE: To assess the validity and reliability of the Korean version of the Virtual Patient Learning System Evaluation Tool (K-VPLSET). DESIGN: This is a methodological study. SETTINGS: This study was conducted via an online survey for Korean nursing students. PARTICIPANTS: The present study included 373 participants who were 3rd and 4th year Korean nursing students. METHODS: After translating the English version of VPLSET into Korean, a pilot test with a cognitive interview was undertaken to ensure that the meaning of original instrument and appropriateness for Korean nursing students had been retained. The content validity of the K-VPLSET was examined by a panel of six experts. Convenience sampling was used to recruit 3rd and 4th year Korean nursing students, among whom 170 were recruited for exploratory factor analysis (EFA) and 203 for confirmatory factor analysis (CFA). SPSS version 26.0 was used for EFA, whereas AMOS version 22.0 was used for CFA. RESULTS: From the 32 initial items, the final version of the K-VPLSET ultimately included 20 items, with a Cronbach's α of 0.89. EFA identified four factors ("Nursing Knowledge Improvement," "Clinical Competency Development," "Confidence in Nursing Performance," and "Nursing Care Plan Application") that explained 56.9% of the total variance. CFA confirmed the validity of the instrument. CONCLUSIONS: Our findings confirmed that the K-VPLSET is a valid and reliable instrument for assessing the effects of the virtual patient learning system, through which the quality of e-learning for Korea nursing students can be determined.

Dietary compound ellagic acid alleviates skin wrinkle and inflammation induced by UV-B irradiation.

Ellagic acid, a polyphenol compound present in berries and pomegranate, has received attention as an agent that may have potential bioactivities preventing chronic diseases. This study examined photoprotective effects of ellagic acid on collagen breakdown and inflammatory responses in UV (ultraviolet)-B irradiated human skin cells and hairless mice. Ellagic acid attenuated the UV-B-induced toxicity of HaCaT keratinocytes and human dermal fibroblasts. Non-toxic ellagic acid markedly prevented collagen degradation by blocking matrix metalloproteinase production in UV-B-exposed fibroblasts. Anti-wrinkle activity of ellagic acid was further investigated in hairless mice exposed to UV-B, in which it attenuated UV-B-triggered skin wrinkle formation and epidermal thickening. Topical application of 10 micromol/l ellagic acid diminished production of pro-inflammatory cytokines IL-1beta and IL-6, and blocked infiltration of inflammatory macrophages in the integuments of SKH-1 hairless mice exposed to UV-B for 8 weeks. In addition, this compound mitigated inflammatory intracellular cell adhesion molecule-1 expression in UV-B-irradiated keratinocytes and photoaged mouse epidermis. These results demonstrate that ellagic acid prevented collagen destruction and inflammatory responses caused by UV-B. Therefore, dietary and pharmacological interventions with berries rich in ellagic acid may be promising treatment strategies interrupting skin wrinkle and inflammation associated with chronic UV exposure leading to photoageing.

Impact of Sarcopenia on Early Postoperative Complications in Early-Stage Non-Small-Cell Lung Cancer.

BACKGROUND: Risk assessment for pulmonary resection in patients with early-stage non-small-cell lung cancer (NSCLC) is important for minimizing postoperative morbidity. Depletion of skeletal muscle mass is closely associated with impaired nutritional status and limited physical ability. We evaluated the relationship between skeletal muscle depletion and early postoperative complications in patients with early-stage NSCLC. METHODS: Patients who underwent curative lung resection between 2016 and 2018 and who were diagnosed with pathological stage I/II NSCLC were included, and their records were retrospectively analyzed. The psoas volume index (PVI, cm3/m3) was calculated based on computed tomography images from routine preoperative positron emission tomography-computed tomography. Early postoperative complications, defined as those occurring within 90 days of surgery, were compared between the lowest sex-specific quartile for PVI and the remaining quartiles. RESULTS: A strong correlation was found between the volume and the cross-sectional area of the psoas muscle (R2=0.816). The overall rate of complications was 57.6% among patients with a low PVI and 32.8% among those with a normal-to-high PVI. The most common complication was prolonged air leak (low PVI, 16.9%; normal-to-high PVI, 9.6%), followed by pneumonia (low PVI, 13.6%; normal-to-high PVI, 7.9%) and recurrent pleural effusion (low PVI, 11.9%; normal-to-high PVI, 6.8%). The predictors of overall complications were low PVI (odds ratio [OR], 2.18; 95% confidence interval [CI], 1.07-4.09; p=0.03), low hemoglobin level (OR, 0.686; 95% CI, 0.54-0.87; p=0.002), and smoking history (OR, 3.93; 95% CI, 2.03-7.58; p<0.001). CONCLUSION: Low PVI was associated with a higher rate of early postoperative complications in patients with early-stage NSCLC.

Nodal Outcomes of Uniportal versus Multiportal Video-Assisted Thoracoscopic Surgery for Clinical Stage I Lung Cancer.

BACKGROUND: Accurate intraoperative assessment of mediastinal lymph nodes is a critical aspect of lung cancer surgery. The efficacy and potential for upstaging implicit in these dissections must therefore be revisited in the current era of uniportal video-assisted thoracoscopic surgery (VATS). METHODS: A retrospective study was conducted in which 544 patients with stage I (T1abc-T2a, N0, M0) primary lung cancer were analyzed. To assess risk factors for nodal upstaging and to limit any imbalance imposed by surgical choices, we constructed an inverse probability of treatment-weighted (IPTW) logistic regression model (in addition to non-weighted logistic models). We also evaluated risk factors for early locoregional recurrence using IPTW logistic regression analysis. RESULTS: In the comparison of uniportal and multiportal VATS, the resected lymph node count (14.03±8.02 vs. 14.41±7.41, respectively; p=0.48) and rate of nodal upstaging (6.5% vs. 8.7%, respectively; p=0.51) appeared similar. Predictors of nodal upstaging included tumor size (odds ratio [OR], 1.74; 95% confidence interval [CI], 1.12-2.70), carcinoembryonic antigen level (OR, 1.11; 95% CI, 1.04-1.18), and histologically confirmed pleural invasion (OR, 3.97; 95% CI, 1.89-8.34). The risk factors for locoregional recurrence within 1 year were found to be number of resected N2 nodes, age, and nodal upstaging. CONCLUSION: Uniportal and multiportal VATS appear similar with regard to accuracy and thoroughness, showing no significant difference in the extent of nodal dissection.

Artificial Nucleosides as Diagnostic Probes to Measure Translesion DNA Synthesis.

The misreplication of damaged DNA, a biological process termed translesion DNA synthesis (TLS), produces a large number of adverse effects on human health. This chapter describes the application of an artificial nucleoside/nucleotide system that functions as a biochemical probe to quantify TLS activity under in vitro and in vivo conditions. For in vitro studies, the artificial nucleotide, 3-ethynyl-5-nitroindolyl-2'-deoxyriboside triphosphate (3-Eth-5-NITP), is used as it is efficiently inserted opposite an abasic site, a highly pro-mutagenic DNA lesion produced by several types of DNA-damaging agents. The placement of the ethynyl moiety allows the incorporated nucleoside triphosphate to be selectively tagged with azide-containing fluorophores via "click" chemistry. This reaction provides a facile way to quantify the extent of nucleotide incorporation opposite this and other noninstructional DNA lesions. The corresponding nucleoside, 3-Eth-5-NIdR, can be used to monitor TLS activity in hematological and adherent cancer cells treated with compounds that produce noninstructional DNA lesions. As described above, visualizing the replication of these lesions is achieved using copper-catalyzed "click" chemistry to tag the ethynyl moiety present on the nucleotide with fluorogenic probes. This technique represents a new diagnostic approach to quantify TLS activity inside cells. In addition, the application of this "clickable" nucleoside provides a chemical probe to identify cells that become drug resistant by the facile replication of noninstructional DNA lesions produced by DNA-damaging agents.