Novel Processes Associated with the Repair of Interstrand Cross-Links Derived from Abasic Sites in Duplex DNA: Roles for the Base Excision Repair Glycosylase NEIL3 and the SRAP Protein HMCES.

Recent studies have defined a novel pathway for the repair of interstrand cross-links derived from the reaction of an adenine residue with an apurinic/apyrimidinic (AP) site on the opposing strand of DNA (dA-AP ICL). Stalling of a replication fork at the dA-AP ICL triggers TRAIP-dependent ubiquitylation of the CMG helicase that recruits the base excision repair glycosylase NEIL3 to the lesion. NEIL3 unhooks the dA-AP ICL to regenerate the native adenine residue on one strand and an AP site on the other strand. Covalent capture of the abasic site by the SRAP protein HMCES protects against genomic instability that would result from cleavage of the abasic site in the context of single-stranded DNA at the replication fork. After repair synthesis moves the HMCES-AP adduct into the context of double-stranded DNA, the DNA-protein cross-link is resolved by a nonproteolytic mechanism involving dissociation of thiazolidine attachment. The AP site in duplex DNA is then repaired by the base excision repair pathway.

Ultrafast Reaction of the Drug Hydralazine with Apurinic/Apyrimidinic Sites in DNA Gives Rise to a Stable Triazolo[3,4-a]phthalazine Adduct.

The clinically used antihypertensive agent hydralazine rapidly generates hydrazone-derived adducts by reaction with apurinic/apyrimidinic (also known as abasic or AP) sites in many different sequences of duplex DNA. The reaction rates are comparable to those of some AP-trapping reagents previously described as "ultrafast." Initially, reversible formation of a hydrazone adduct is followed by an oxidative cyclization reaction that generates a chemically stable triazolo[3,4-a]phthalazine adduct. The net result is that the reaction of hydralazine with AP sites in duplex DNA yields a rapid and irreversible adduct formation. Although the hydrazone and triazolo[3,4-a]phthalazine adducts differ by only two mass units, it was possible to use MALDI-TOF-MS and ESI-QTOF-nanospray-MS to quantitatively characterize mixtures of these adducts by deconvolution of overlapping isotope envelopes. Reactions of hydralazine with the endogenous ketone pyruvate do not prevent the formation of the hydralazine-AP adducts, providing further evidence that these adducts have the potential to form in cellular DNA. AP sites are ubiquitous in cellular DNA, and rapid, irreversible adduct formation by hydralazine could be relevant to the pathogenesis of systemic drug-induced lupus erythematosus experienced by some patients. Finally, hydralazine might be developed as a probe for the detection of AP sites, the study of cellular BER, and marking the location of AP sites in DNA-sequencing analyses.

Formation and Properties of DNA Adducts Generated by Reactions of Abasic Sites with 1,2-Aminothiols Including Cysteamine, Cysteine Methyl Ester, and Peptides Containing N-Terminal Cysteine Residues.

The reaction of 1,2-aminothiol groups with aldehyde residues in aqueous solution generates thiazolidine products, and this process has been developed as a catalyst-free click reaction for bioconjugation. The work reported here characterized reactions of the biologically relevant 1,2-aminothiols including cysteamine, cysteine methyl ester, and peptides containing N-terminal cysteine residues with the aldehyde residue of apurinic/apyrimidinic (AP) sites in DNA oligomers. These 1,2-aminothiol-containing compounds rapidly generated adducts with AP sites in single-stranded and double-stranded DNA. NMR and MALDI-TOF-MS analyses provided evidence that the reaction generated a thiazolidine product. Conversion of an AP site to a thiazolidine-AP adduct protected against the rapid cleavage normally induced at AP sites by the endonuclease action of the enzyme APE1 and the AP-lyase activity of the biogenic amine spermine. In the presence of excess 1,2-aminothiols, the thiazolidine-AP adducts underwent slow strand cleavage via a ß-lyase reaction that generated products with 1,2-aminothiol-modified sugar residues on the 3'-end of the strand break. In the absence of excess 1,2-aminothiols, the thiazolidine-AP adducts dissociated to release the parent AP-containing oligonucleotide. The properties of the thiazolidine-AP adducts described here mirror critical properties of SRAP proteins HMCES and YedK that capture AP sites in single-stranded regions of cellular DNA and protect them from cleavage.

Severe inframalleolar disease is an independent predictor of adverse limb outcomes after endovascular revascularization in chronic limb-threatening ischemia.

OBJECTIVE: In the present study, we evaluated the effects of inframalleolar (IM) disease on the occurrence of major adverse limb events (MALE) in patients undergoing endovascular revascularization for chronic limb-threatening ischemia (CLTI). METHODS: Patients who had undergone endovascular revascularization for CLTI between January 2015 and December 2019 at two university-affiliated hospitals were reviewed retrospectively. Patients with severe IM disease (pedal score of 2) were compared with those with mild to moderate IM disease (score of 0 or 1) using the Global Vascular Guidelines. The primary outcome was MALE (open revascularization, acute leg ischemia, major amputation). The secondary outcomes were mortality, reintervention, major adverse cardiac events, and perioperative complications ≤30 days after endovascular revascularization, primary limb-based patency, and the occurrence of any limb event (defined as any amputation, acute leg ischemia, or open revascularization). Kaplan-Meier estimates were used to compare the primary outcome, and the Cox proportion hazard model was used to assess the effects of IM disease. RESULTS: The study included 167 limbs in 149 patients (36% female; mean age, 74 ± 12 years). Severe IM disease was identified in 71 limbs (43%). No differences were found in the baseline characteristics, except for a higher prevalence of dyslipidemia in the patients with severe IM disease (66% vs 43%; P = .003). Most patients in both groups had had a WIfI (Wound, Ischemia, foot Infection) score of 4 (severe IM disease, 64%; vs mild to moderate IM disease, 57%; P = .462) and GLASS (global limb anatomic severity scale) III anatomy (severe IM disease, 54%; vs mild to moderate IM disease, 48%; P = .752). The Kaplan-Meier estimates showed that severe IM disease was associated with lower freedom from MALE (69% vs 82%; P = .026). The Cox proportion hazard regression model showed that severe IM disease was an independent predictor of increased MALE and amputation risk (hazard ratio, 1.715; 95% confidence interval, 1.015-2.896; P = .044) after adjusting for covariates. During follow-up, patients with severe IM disease had had mortality (27% vs 31%; P = .567) and reintervention (42% vs 38%; P = .608) similar to those for patients with mild to moderate IM disease. Primary limb-based patency was also similar (79% vs 84%; P = .593) at a mean follow-up of 3.8 ± 0.8 years. CONCLUSIONS: Severe IM disease was prevalent in 43% of limbs that had undergone endovascular revascularization for CLTI and was associated with lower freedom from MALE. Severe IM disease also independently increased the hazard of adverse limb outcomes and amputations in patients with CLTI by >70%, highlighting its importance as a measure of foot perfusion.

A systematic review and meta-analysis of the effect of obesity on patients undergoing lower extremity revascularization.

OBJECTIVE: In the present review, we assessed the effect of obesity on clinical outcomes for patients with peripheral arterial disease who had undergone endovascular or open lower extremity revascularization surgery. METHODS: A systematic search strategy of MEDLINE, EMBASE, CINAHL, Web of Science, and Cochrane Library was conducted. The included studies had compared obese and nonobese cohorts with peripheral arterial disease who had undergone endovascular or open lower extremity revascularization. The outcomes included mortality, major adverse cardiovascular events, major adverse limb events, surgical site infections, endovascular access site complications, and perioperative complications. RESULTS: Eight studies were included with 171,648 patients. The obese patients (body mass index ≥30 kg/m2) were more likely to be women, to have diabetes, and to have more cardiovascular comorbidities despite being younger. No association was found between obesity and peripheral arterial disease severity. Obesity was associated with an overall 22% decreased mortality risk after lower extremity revascularization (risk ratio [RR], 0.78; 95% confidence interval [CI], 0.71-0.85; P < .001; I2 = 0%; GRADE (grading of recommendations assessment, development, evaluation), very low quality). A subgroup analysis by intervention type showed similar findings (endovascular: RR, 0.79; 95% CI, 0.71-0.87; P < .001; I2 = 0%; open: RR, 0.70; 95% CI, 0.51-0.95; P = .024; I2 = 43%). Obesity was associated with a 14% decreased risk of major adverse cardiovascular events for open surgery only (RR, 0.86; 95% CI, 0.76-0.98; P = .021; I2 = 0%; GRADE, very low quality). Obesity was associated with an increased risk of surgical site infections pooled across intervention types (RR, 1.69; 95% CI, 1.34-2.14; P < .001; I2 = 78%; GRADE, very low quality). No association was found between obesity and major adverse limb events (RR, 1.02; 95% CI, 0.93-1.11; P = .73; I2 = 15%; GRADE, very low quality) or endovascular access site complications (RR, 1.11; 95% CI, 0.76-1.63; P = .58; I2 = 86%; GRADE, very low quality). Pooled perioperative complications did not differ between the obese and nonobese cohorts (RR, 1.04; 95% CI, 0.84-1.28; P = .73; I2 = 92%; GRADE, very low quality). CONCLUSIONS: Obesity was associated with reduced mortality risk with both endovascular and open surgery, although a reduction in major adverse cardiovascular events was only observed with open surgery. In addition, obese patients had an increased risk of surgical site infections. Obesity was not associated with major adverse limb events, endovascular access site complications, or perioperative complications. The GRADE quality of evidence was very low. The findings from the present review suggest a survival advantage for obese patients with peripheral arterial disease. Future studies could focus on prospectively investigating the effect of obesity on peripheral arterial disease outcomes. A nuanced evaluation of body mass index as a preoperative risk factor is warranted.

Multimodal Prehabilitation for Patients Undergoing Endovascular Aortic Aneurysm Repair Surgery: A Feasibility Study.

BACKGROUND: Elective treatment options for aortic abdominal aneurysms include open repair or the less-invasive endovascular aortic aneurysm repair (EVAR). Recovery from EVAR is generally considered easier and faster than open repair. Despite this, EVAR remains a major procedure, and average return to preoperative quality of life is at least 3 months. The purpose of this study is to determine the safety and feasibility of multimodal prehabilitation, a multidisciplinary preoperative optimization intervention, in patients undergoing EVAR and its impact on perioperative functional capacity and quality of life. METHODS: Candidates for EVAR with an infra-renal abdominal aortic aneurysm <7.5cm were invited to participate in a 6-week multimodal prehabilitation program that included (1) supervised and home-based exercise, (2) nutritional support, (3) psychosocial support, and (4) smoking cessation. Functional capacity and quality of life were assessed at baseline, before surgery and 6 weeks postoperatively. Recruitment rate, safety, and compliance were also assessed. RESULTS: A total of 24 patients were included, 17 males (70%) and 7 females (30%). No adverse events occurred during the program. Compliance to each component of the program (median [Q1-Q3]) was 66% [67] for supervised training, 100% [67] for home-based training, and 100% [100] for nutrition. The multimodal prehabilitation program elicited a significant increase in functional capacity and quality of life preoperatively. CONCLUSION: Multimodal prehabilitation for patients awaiting EVAR is feasible and safe. Multimodal prehabilitation improves both functional capacity and quality of life preoperatively. Further research is needed to assess the impact of multimodal prehabilitation on postoperative quality of life and functional capacity. CLINICAL IMPACT: Multimodal prehabilitation is safe and feasible in patients awaiting endovascular aneurysm repair. The importance of this finding is that multimodal prehabilitation can be safely delivered preoperatively in patients awaiting EVAR. Although further research is needed, multimodal prehabilitation seems to improve preoperative functional capacity and quality of life. This could have an impact for the future implementation of prehabilitation interventions in order to increase functional reserve and quality of life preoperatively so that this high-risk population can cope better with the surgical stress and return to their normal life faster postoperatively.

Association Between Obesity and Outcomes Following Endovascular Aneurysm Repair.

BACKGROUND: Obesity is prevalent in patients with abdominal aortic aneurysms (AAA). There is an association between increasing body mass index (BMI) and increased overall cardiovascular mortality and morbidity. This study aims to assess the difference in mortality and complication rates between normal weight (NW), overweight (OW), and obese patients undergoing endovascular aneurysm repair (EVAR) for infrarenal AAA. METHODS: This is a retrospective analysis of consecutive patients undergoing EVAR for AAA between January 1998 and December 2019. Weight classes were defined as: BMI<18.5 kg/m2, underweight; BMI 18.5-24.9 kg/m2, NW; BMI 25.0-29.9 kg/m2, OW; BMI 30.0-39.9 kg/m2, obese; BMI>39.9 kg/m2 morbidly obese. Primary outcomes were long-term all-cause mortality and freedom from reintervention. Secondary outcome was aneurysm sac regression (defined as a reduction in sac diameter of 5 mm or more). Kaplan-Meier survival estimates and mixed model analysis of variance were used. RESULTS: The study included 515 patients (83% males, mean age 77 ± 8 years) with a mean follow-up of 3.8 ± 2.8 years. In terms of weight class, 2.1% (n = 11) were underweight, 32.4% (167) were NW, 41.6% (n = 214) were OW, 21.2% (n = 109) were obese, and 2.7% (n = 14) were morbidly obese. Obese patients were younger (mean difference -5.0 years) but had a higher prevalence of diabetes mellitus (33.3% vs. 10.6% for NW) and dyslipidemia (82.4% vs. 60.9% for NW). Obese patients had similar freedom from all-cause mortality (88%) compared to OW (78%) and NW (81%) patients. The same findings were evident for freedom from reintervention where obese (79%) was similar to OW (76%) and NW (79%). At a mean follow-up of 5.1 ± 0.4 years, sac regression was observed similarly across weight classes at 49.6%, 50.6%, and 51.8% for NW, OW, and obese, respectively (P = 0.501). There was a significant difference in mean AAA diameter pre- and post-EVAR [F(2,318) = 24.37, P < 0.001] across weight classes. NW [mean reduction 4.8 mm (2.0-7.6 mm, P < 0.001)], OW [mean reduction 3.9 mm (1.5-6.3 mm, P < 0.001)], and obese [mean reduction 5.7 mm (2.3-9.1 mm, P < 0.001)] achieved similar reductions. CONCLUSIONS: Obesity was not associated with increased mortality or reintervention in patients undergoing EVAR. Obese patients achieved similar rates of sac regression on imaging follow-up.

Multimodal Prehabilitation for Peripheral Arterial Disease Patients with Intermittent Claudication-A Pilot Randomized Controlled Trial.

BACKGROUND: To establish the feasibility and safety of multimodal prehabilitation (MP), and to obtain pilot data on the change in quality of life, functional walking capacity, and the need for surgery for a full-scale trial. METHODS: Pilot randomized controlled trial that included patients older than 50 years old suffering from moderate to severe intermittent claudication and who were candidates for endovascular revascularization (ER). Participants were excluded if they presented with ischemic rest pain, gangrene or ulceration of the index leg, significant lesions in the iliac vessels, planned surgical bypass, comorbidities in which exercise was contraindicated or if they were unable to speak English or French. Participants were randomized in a 1:1 ratio to 12 weeks of MP or institutional standard of care (unsupervised walking advice). MP consisted of i)1 weekly supervised exercise session; ii) home-based exercise prescription; iii) nutritional counseling and supplementation; iv) smoking cessation therapy; and v) psychosocial support. Feasibility and safety were measured with recruitment and retention rates, as well as the occurrence of any adverse events. In addition, barriers to attend supervised sessions and compliance to each component were assessed. Change in functional walking capacity, health-related quality of life, and the rates of patients deciding not to undergo ER were collected and analyzed throughout the follow-up period of 12 months. RESULTS: Of the 37 patients referred for eligibility, 27 (73%) accepted to participate in the trial and were randomized. Of the 27 patients included, 24 completed the 12-week program. Adherence to each prehabilitation component was 83% interquartile range [72,93] for supervised exercise, 90% [83,96] for home-based exercise and 69% [45,93] for nutritional sessions. Fifty percent of patients were referred for and underwent psychosocial intervention and 40% of the active smokers enrolled in the smoking cessation program. No adverse events were observed during the program. The 2 main barriers for not fully adhering to the intervention were excessive pain while performing the exercises and the difficulty to keep up with the prescribed exercises. A statistically significant mean change (standard deviation (SD)) was seen in the MP group versus standard of care for functional capacity, mean (SD) 6 Min Walk Test 60 (74) vs. -11 (40) meters P < 0.05, and quality of life mean (SD) VascuQol 1.15 (0.54) vs. -0.3 (1.09) points P < 0.05. There was no statistically significant difference between groups in the rates of patients deciding to undergo ER during the 1-year follow-up period. CONCLUSIONS: The results of this pilot trial demonstrate that MP is safe and feasible. A 12-week MP program seems to improve quality of life and functional walking capacity to a greater extent than unsupervised walking advice. There is a need for a large-scale trial to investigate the effectiveness of MP at improving quality of life and assessing its impact on the rates of patients deciding not to undergo or delay ER. The long-term functional and quality of life outcomes of the patients deciding to undergo ER after prehabilitation also need to be assessed.

Reconsidering the Chemical Nature of Strand Breaks Derived from Abasic Sites in Cellular DNA: Evidence for 3'-Glutathionylation.

The hydrolytic loss of coding bases from cellular DNA is a common and unavoidable reaction. The resulting abasic sites can undergo ß-elimination of the 3'-phosphoryl group to generate a strand break with an electrophilic α,ß-unsaturated aldehyde residue on the 3'-terminus. The work reported here provides evidence that the thiol residue of the cellular tripeptide glutathione rapidly adds to the alkenal group on the 3'-terminus of an AP-derived strand break. The resulting glutathionylated adduct is the only major cleavage product observed when ß-elimination occurs at an AP site in the presence of glutathione. Formation of the glutathionylated cleavage product is reversible, but in the presence of physiological concentrations of glutathione, the adduct persists for days. Biochemical experiments provided evidence that the 3'-phosphodiesterase activity of the enzyme apurinic/apyrimidinic endonuclease (APE1) can remove the glutathionylated sugar remnant from an AP-derived strand break to generate the 3'OH residue required for repair via base excision or single-strand break repair pathways. The results suggest that a previously unrecognized 3'glutathionylated sugar remnant─and not the canonical α,ß-unsaturated aldehyde end group─may be the true strand cleavage product arising from ß-elimination at an abasic site in cellular DNA. This work introduces the 3'glutathionylated cleavage product as the major blocking group that must be trimmed to enable repair of abasic site-derived strand breaks by the base excision repair or single-strand break repair pathways.

N-Methyl-N-nitrosourea Induced 3'-Glutathionylated DNA-Cleavage Products in Mammalian Cells.

Apurinic/apyrimidinic (AP) sites, that is, abasic sites, are among the most frequently induced DNA lesions. Spontaneous or DNA glycosylase-mediated ß-elimination of the 3'-phosphoryl group can lead to strand cleavages at AP sites to yield a highly reactive, electrophilic 3'-phospho-α,ß-unsaturated aldehyde (3'-PUA) remnant. The latter can react with amine or thiol groups of biological small molecules, DNA, and proteins to yield various damaged 3'-end products. Considering its high intracellular concentration, glutathione (GSH) may conjugate with 3'-PUA to yield 3-glutathionyl-2,3-dideoxyribose (GS-ddR), which may constitute a significant, yet previously unrecognized endogenous lesion. Here, we developed a liquid chromatography tandem mass spectroscopy method, in combination with the use of a stable isotope-labeled internal standard, to quantify GS-ddR in genomic DNA of cultured human cells. Our results revealed the presence of GS-ddR in the DNA of untreated cells, and its level was augmented in cells upon exposure to an alkylating agent, N-methyl-N-nitrosourea (MNU). In addition, inhibition of AP endonuclease (APE1) led to an elevated level of GS-ddR in the DNA of MNU-treated cells. Together, we reported here, for the first time, the presence of appreciable levels of GS-ddR in cellular DNA, the induction of GS-ddR by a DNA alkylating agent, and the role of APE1 in modulating its level in human cells.

Unexpected Complexity in the Products Arising from NaOH-, Heat-, Amine-, and Glycosylase-Induced Strand Cleavage at an Abasic Site in DNA.

Hydrolytic loss of nucleobases from the deoxyribose backbone of DNA is one of the most common unavoidable types of damage in synthetic and cellular DNA. The reaction generates abasic sites in DNA, and it is important to understand the properties of these lesions. The acidic nature of the α-protons of the ring-opened abasic aldehyde residue facilitates the ß-elimination of the 3'-phosphoryl group. This reaction is expected to generate a DNA strand break with a phosphoryl group on the 5'-terminus and a trans-α,ß-unsaturated aldehyde residue on the 3'-terminus; however, a handful of studies have identified noncanonical sugar remnants on the 3'-terminus, suggesting that the products arising from strand cleavage at apurinic/apyrimidinic sites in DNA may be more complex than commonly thought. We characterized the strand cleavage induced by the treatment of an abasic site-containing DNA oligonucleotide with heat, NaOH, piperidine, spermine, and the base excision repair glycosylases Fpg and Endo III. The results showed that under multiple conditions, cleavage at an abasic site in a DNA oligomer generated noncanonical sugar remnants including cis-α,ß-unsaturated aldehyde, 2-deoxyribose, and 3-thio-2,3-dideoxyribose products on the 3'-terminus of the strand break.

Products Generated by Amine-Catalyzed Strand Cleavage at Apurinic/Apyrimidinic Sites in DNA: New Insights from a Biomimetic Nucleoside Model System.

Abasic sites are common in cellular and synthetic DNA. As a result, it is important to characterize the chemical fate of these lesions. Amine-catalyzed strand cleavage at abasic sites in DNA is an important process in which conversion of small amounts of the ring-opened abasic aldehyde residue to an iminium ion facilitates ß-elimination of the 3'-phosphoryl group. This reaction generates a trans-α,ß-unsaturated iminium ion on the 3'-terminus of the strand break as an obligate intermediate. The canonical product expected from amine-catalyzed cleavage at an AP site is the corresponding trans-α,ß-unsaturated aldehyde sugar remnant resulting from hydrolysis of this iminium ion. Interestingly, a handful of studies have reported noncanonical 3'-sugar remnants generated by amine-catalyzed strand cleavage, but the formation and properties of these products are not well-understood. To address this knowledge gap, a nucleoside system was developed that enabled chemical characterization of the sugar remnants generated by amine-catalyzed ß-elimination in the 2-deoxyribose system. The results predict that amine-catalyzed strand cleavage at an AP site under physiological conditions has the potential to reversibly generate noncanonical cleavage products including cis-alkenal, 3-thio-2,3-dideoxyribose, and 2-deoxyribose groups alongside the canonical trans-alkenal residue on the 3'-terminus of the strand break. Thus, the model reactions provide evidence that the products generated by amine-catalyzed strand cleavage at abasic sites in cellular DNA may be more complex that commonly thought, with trans-α,ß-unsaturated iminium ion intermediates residing at the hub of interconverting product mixtures. The results expand the list of possible 3'-sugar remnants arising from amine-catalyzed cleavage of abasic sites in DNA that must be chemically or enzymatically removed for the completion of base excision repair and single-strand break repair in cells.

Feasibility and Cost Analysis of Ambulatory Endovascular Aneurysm Repair.

PURPOSE: We sought to compare the costs of ambulatory endovascular aneurysm repair (a-EVAR) and inpatient EVAR (i-EVAR) at up to 1-year of follow-up. MATERIALS AND METHODS: A retrospective cohort study of consecutive patients undergoing elective EVAR between April 2016 and December 2018 at two academic centers. Patients planned for a-EVAR were compared with i-EVAR. Costs at 30 days and 1 year were extracted. These included operating room (OR) use, bed occupancy, laboratory and imaging, emergency department (ED) visits, readmissions, and reinterventions. Baseline characteristics were compared. Multiple regression model was used to identify predictors of increased EVAR costs. Repeated measures analysis of variance (ANOVA) was used to compare cost differences at 30 days and 1 year via an intention-to-treat analysis. Bonferroni post hoc test compared between-group differences. A p value<0.05 was considered statistically significant. RESULTS: One hundred seventy patients were included. Most underwent percutaneous EVAR (>94%) under spinal anesthesia (>84%). Ambulatory endovascular aneurysm repair was successful in 84% (84/100). Ambulatory endovascular aneurysm repair patients (76±8 years) were younger than i-EVAR (78±9 years). They also had a smaller mean aneurysm diameter (56±6 mm) compared with i-EVAR (59±6 mm). Emergency department visits, readmissions, and reinterventions were similar up to 1 year (all p=NS). Ambulatory endovascular aneurysm repair costs showed a non-statistically significant reduction in total costs at 30 days and 1 year by 27% and 21%, respectively. Patients younger than 85 years and males had a 30-day cost reduction by 34% (p=0.027) and 33% (p=0.035), respectively with a-EVAR. CONCLUSIONS: Same-day discharge is feasible and successful in selected patients. Patients younger than 85 years and males have a short-term cost benefit with EVAR done in the ambulatory setting without increased complications or reinterventions. CLINICAL IMPACT: This study shows the overall safety of ambulatory EVAR with proper patient selection. These patient had similar post-intervention complications to inpatients. Same day discharge also resulted in short-term reduction in costs in male patients and patients younger than 85 years.

Comorbid Depression is Associated with Increased Major Adverse Limb Events in Peripheral Arterial Disease: A Systematic Review and Meta-analysis.

OBJECTIVE: Depression is a significant risk factor for death in coronary artery disease. Conversely, the research surrounding depression and peripheral arterial disease is limited. This review aimed to systematically evaluate the available literature on the impact of comorbid depression on adverse outcomes in peripheral arterial disease. DATA SOURCES: A systematic review and meta-analysis were performed using the following databases MEDLINE, EMBASE, CINAHL, PsycINFO, and Cochrane Library from inception until July 2021. REVIEW METHODS: Included studies compared depressed and non-depressed patients with peripheral arterial disease. The outcomes included death, major adverse cardiovascular events, and major adverse limb events. RESULTS: A total of 9 297 articles were searched. Of these, seven studies were identified. Depressed patients were more likely to be women, diabetic, have a history of smoking, and have chronic limb threatening ischaemia, despite being younger than non-depressed patients. There was a 20% increase in major adverse limb events in depressed patients (RR 1.20, 95% CI 1.11 - 1.31, z = 3.9, p < .001, GRADE strength: very low) but no increased risk of death (RR 1.03, 95% CI 0.72 - 1.40, z = 0.06, p = .95, GRADE strength: very low) or major adverse cardiovascular events (RR 1.16, 95% CI 0.67 - 2.01, z = 0.54, p = .59, GRADE strength: very low). A follow up meta-regression of various comorbidities and demographic variables did not demonstrate a significant contribution to the observed risk ratio for major adverse limb events. CONCLUSION: Depression was reported in 13% of patients with peripheral arterial disease, associated with more medical comorbidity, and a 20% increased risk of major adverse limb events. Although the strength of this evidence is very low, the current state of the literature remains limited. Future studies should prospectively assess the impact of depression and its relationship to medical comorbidities and high risk health behaviours.

Outcomes of endovascular treatment of patients with intermittent claudication due to femoropopliteal disease.

OBJECTIVE: Our objective was to evaluate the outcomes of endovascular treatment in patients with moderate and severe claudication due to femoropopliteal disease, that is, disease of the superficial femoral and popliteal arteries. METHODS: A retrospective review of all patients with moderate and severe claudication (Rutherford 2 and 3) undergoing endovascular treatment for FP disease between January 2012 and December 2017 at two university-affiliated hospitals was performed. All procedures were performed by vascular surgeons. Primary outcomes were mortality, freedom from reintervention, major adverse limb events defined as major amputations, open surgical revascularization, or progression to chronic limb-threatening ischemia (CLTI) at 30 days, 1 year, 2 years, and last follow-up. Unadjusted odds ratios were calculated to identify variables associated with adverse outcomes, and Kaplan-Meier survival curves were used to determine mortality and freedom from reintervention. RESULTS: Eighty-five limbs in 74 patients were identified on review. Mean age was 69.6 ± 9.8 years and 74.3% were males. At a median follow-up of 49.0 ± 25.5 months, all-cause mortality rate was 8.1% (6 patients) with 16.7% being due to cardiovascular causes. Reintervention rates were 1.2%, 16.5%, and 21.2% at 30 days, 1 year, and 2 years, respectively. Major adverse limb events occurred in 3 patients and rates were 0%, 1.2%, and 2.4% at 30 days, 1 year, and 2 years, respectively. Progression to CLTI was 0%, 1.2%, and 1.2% at 30 days, 1 year, and 2 years, respectively. Claudication had improved or resolved in 55.6% (n = 34 patients), stable in 38.9% (n = 21 patients), and worse in 5.6% (n = 3 patients) Age ≥ 70 years (OR = 4.09 (1.14-14.66), p = 0.027), TASCII A lesion (OR = 4.67 (1.14-19.17), p = 0.025), and presence of 3-vessel runoff (OR = 3.70 (1.18-11.59), p = 0.022) predicted symptoms' improvement. TASCII A lesions were less likely to require reintervention (OR = 0.23 (0.06-0.86), p = 0.020). Reintervention within 1 year (OR = 11.67 (0.98-138.94), p = 0.017), reintervention with a stent (OR = 14.40 (1.19-173.67), p = 0.008) and more than one reintervention (OR = 39.00 (2.89-526.28), p < 0.001) predicted major adverse limb events. CONCLUSIONS: Careful patient selection is important when planning endovascular treatment in patients with intermittent claudication and FP disease. This could result in symptomatic improvement in more than half of the patients. Adverse outcomes such as major adverse limb events, progression to CLTI, and amputations occur at low rates.

Structure of a Stable Interstrand DNA Cross-Link Involving a ß-N-Glycosyl Linkage Between an N6-dA Amino Group and an Abasic Site.

Abasic (AP) sites are one of the most common forms of DNA damage. The deoxyribose ring of AP sites undergoes anomerization between α and ß configurations, via an electrophilic aldehyde intermediate. In sequences where an adenine residue is located on the opposing strand and offset 1 nt to the 3' side of the AP site, the nucleophilic N6-dA amino group can react with the AP aldehyde residue to form an interstrand cross-link (ICL). Here, we present an experimentally determined structure of the dA-AP ICL by NMR spectroscopy. The ICL was constructed in the oligodeoxynucleotide 5'-d(T1A2T3G4T5C6T7A8A9G10T11T12C13A14T15C16T17A18)-3':5'-d(T19A20G21A22T23G24A25A26C27X28T29A30G31A32C33A34T35A36)-3' (X=AP site), with the dA-AP ICL forming between A8 and X28. The NMR spectra indicated an ordered structure for the cross-linked DNA duplex and afforded detailed spectroscopic resonance assignments. Structural refinement, using molecular dynamics calculations restrained by NOE data (rMD), revealed the structure of the ICL. In the dA-AP ICL, the 2'-deoxyribosyl ring of the AP site was ring-closed and in the ß configuration. Juxtapositioning the N6-dA amino group and the aldehydic C1 of the AP site within bonding distance while simultaneously maintaining two flanking unpaired A9 and T29 bases stacked within the DNA is accomplished by the unwinding of the DNA at the ICL. The structural data is discussed in the context of recent studies describing the replication-dependent unhooking of the dA-AP ICL by the base excision repair glycosylase NEIL3.

An autoinhibitory role for the GRF zinc finger domain of DNA glycosylase NEIL3.

The NEIL3 DNA glycosylase maintains genome integrity during replication by excising oxidized bases from single-stranded DNA (ssDNA) and unhooking interstrand cross-links (ICLs) at fork structures. In addition to its N-terminal catalytic glycosylase domain, NEIL3 contains two tandem C-terminal GRF-type zinc fingers that are absent in the other NEIL paralogs. ssDNA binding by the GRF-ZF motifs helps recruit NEIL3 to replication forks converged at an ICL, but the nature of DNA binding and the effect of the GRF-ZF domain on catalysis of base excision and ICL unhooking is unknown. Here, we show that the tandem GRF-ZFs of NEIL3 provide affinity and specificity for DNA that is greater than each individual motif alone. The crystal structure of the GRF domain shows that the tandem ZF motifs adopt a flexible head-to-tail configuration well-suited for binding to multiple ssDNA conformations. Functionally, we establish that the NEIL3 GRF domain inhibits glycosylase activity against monoadducts and ICLs. This autoinhibitory activity contrasts GRF-ZF domains of other DNA-processing enzymes, which typically use ssDNA binding to enhance catalytic activity, and suggests that the C-terminal region of NEIL3 is involved in both DNA damage recruitment and enzymatic regulation.

Formation and Repair of an Interstrand DNA Cross-Link Arising from a Common Endogenous Lesion.

Interstrand DNA cross-links (ICLs) are cytotoxic because they block the strand separation required for read-out and replication of the genetic information in duplex DNA. The unavoidable formation of ICLs in cellular DNA may contribute to aging, neurodegeneration, and cancer. Here, we describe the formation and properties of a structurally complex ICL derived from an apurinic/apyrimidinic (AP) site, which is one of the most common endogenous lesions in cellular DNA. The results characterize a cross-link arising from aza-Michael addition of the N2-amino group of a guanine residue to the electrophilic sugar remnant generated by spermine-mediated strand cleavage at an AP site in duplex DNA. An α,ß-unsaturated iminium ion is the critical intermediate involved in ICL formation. Studies employing the bacteriophage φ29 polymerase provided evidence that this ICL can block critical DNA transactions that require strand separation. The results of biochemical studies suggest that this complex strand break/ICL might be repaired by a simple mechanism in which the 3'-exonuclease action of the enzyme apurinic/apyrimidinic endonuclease (APE1) unhooks the cross-link to initiate repair via the single-strand break repair pathway.

Host resistance to Bacillus thuringiensis is linked to altered bacterial community within a specialist insect herbivore.

Evolution of resistance to transgenic crops producing toxins from Bacillus thuringiensis (Bt) threatens the sustainability of the technology. Examination of resistance mechanisms has largely focused on characterization of mutations in proteins serving as Bt toxin binding sites. However, insect microbial communities have the potential to provide host resistance to pesticides in a myriad of ways. Previous findings suggest the killing mechanism of Bt relies on enteric bacteria becoming pathogenic in the disrupted gut environment of the insect following Bt intoxication. Thus, here we hypothesized that resistance to Bt would alter the microbiome composition of the insect. Previous studies have manipulated the microbiome of susceptible insects and monitored their response to Bt. In our study, we characterized the associated bacterial communities of Bt-resistant and -susceptible western corn rootworms, a widespread pest of maize in the United States. We found resistant insects harbor a bacterial community that is less rich and distinct from susceptible insects. After feeding on Bt-expressing maize, susceptible insects exhibited dysbiosis of the associated bacterial community, whereas the community within resistant insects remained relatively unchanged. These results suggest resistance to Bt produces alterations in the microbiome of the western corn rootworm that may contribute to resistance. We further demonstrated that by itself, feeding on Bt toxin-expressing seedlings caused a shift in the microbiota. This work provides a broader picture of the effect stressors have on microbiome composition, and the potential heritable changes induced as a result of intense selection.

Synthesis of DNA Duplexes Containing Site-Specific Interstrand Cross-Links via Sequential Reductive Amination Reactions Involving Diamine Linkers and Abasic Sites on Complementary Oligodeoxynucleotides.

Interstrand DNA cross-links are important in biology, medicinal chemistry, and materials science. Accordingly, methods for the targeted installation of interstrand cross-links in DNA duplexes may be useful in diverse fields. Here, a simple procedure is reported for the preparation of DNA duplexes containing site-specific, chemically defined interstrand cross-links. The approach involves sequential reductive amination reactions between diamine linkers and two abasic (apurinic/apyrimidinic, AP) sites on complementary oligodeoxynucleotides. Use of the symmetrical triamine, tris(2-aminoethyl)amine, in this reaction sequence enabled the preparation of a cross-linked DNA duplex bearing a derivatizable aminoethyl group.