NIR responsive scaffold with multistep shape memory and photothermal-chemodynamic properties for complex tissue defects repair and antibacterial therapy.

Complex tissue damage accompanying with bacterial infection challenges healthcare systems globally. Conventional tissue engineering scaffolds normally generate secondary implantation trauma, mismatched regeneration and infection risks. Herein, we developed an easily implanted scaffold with multistep shape memory and photothermal-chemodynamic properties to exactly match repair requirements of each part from the tissue defect by adjusting its morphology as needed meanwhile inhibiting bacterial infection on demand. Specifically, a thermal-induced shape memory scaffold was prepared using hydroxyethyl methacrylate and polyethylene glycol diacrylate, which was further combined with the photothermal agent iron tannate (FeTA) to produce NIR light-induced shape memory property. By varying ingredients ratios in each segment, this scaffold could perform a stepwise recovery under different NIR periods. This process facilitated implantation after shape fixing to avoid trauma caused by conventional methods and gradually filled irregular defects under NIR to perform suitable tissue regeneration. Moreover, FeTA also catalyzed Fenton reaction at bacterial infections with abundant H2O2, which produced excess ROS for chemodynamic antibacterial therapy. As expected, bacteriostatic rate was further enhanced by additional photothermal therapy under NIR. The in vitro and vivo results showed that our scaffold was able to perform high efficacy in both antibiosis, inflammation reduction and wound healing acceleration, indicating a promising candidate for the regeneration of complex tissue damage with bacterial infection.

Anti-inflammatory and Restorative effects of milk exosomes and Dexamethasone-Loaded exosomes in a corneal alkali burn model.

Corneal alkali burn is a common and challenging ocular trauma, necessitating the use of dexamethasone (DXMS) as a therapeutic agent. However, prolonged and frequent administration of this drug can lead to undesirable side effects, limiting its clinical application. This study aimed to investigate the role and mechanism of action of exosomes as drug carriers in corneal alkali burn repair. We employed centrifugation to isolate milk exosomes (EXO) as nanocarriers. We observed that EXO enhanced the activity and migration of corneal epithelial cells, expediting the repair process following corneal injury. Additionally, a nano-drug delivery model (DXMS@EXO) was designed using ultrasound to load DXMS into exosomes, thus enabling targeted delivery to inflammatory cells and enhancing drug efficacy. DXMS@EXO inhibited the inflammatory processes in the corneal alkali burn model by modulating the classical Wnt signaling pathway, thereby promoting corneal re-epithelialization and wound healing and accelerating the repair process of corneal alkali burn. Neither EXO nor DXMS@EXO exhibited significant side effects during the course of treatment. This study highlighted the substantial potential of EXO and DXMS@EXO in improving drug efficacy and facilitating the repair of corneal alkali burn.

Role of RNA polymerase III transcription and regulation in ischaemic stroke.

Ischaemic stroke is a leading cause of death and life-long disability due to neuronal cell death resulting from interruption of glucose and oxygen supplies. RNA polymerase III (Pol III)-dependent transcription plays a central role in protein synthesis that is necessary for normal cerebral neuronal functions, and the survival and recovery under pathological conditions. Notably, Pol III transcription is highly sensitive to ischaemic stress that is known to rapidly shut down Pol III transcriptional activity. However, its precise role in ischaemic stroke, especially during the acute and recovery phases, remains poorly understood. The microenvironment within the ischaemic brain undergoes dynamic changes in different phases after stroke. Emerging evidence highlights the distinct roles of Pol III transcription in neuroprotection during the acute phase and repair during the recovery phase of stroke. Additionally, investigations into the mTOR-MAF1 signalling pathway, a conserved regulator of Pol-III transcription, reveal its therapeutic potential in enhancing acute phase neuroprotection and recovery phase repair.

Transcatheter Mitral Valve Repair for Multiple Valvular Heart Disease: Outcomes and Insights on Combined Aortic Insufficiency and Mitral Regurgitation.

The presence of concomitant aortic insufficiency (AI) and mitral regurgitation (MR) is common and may further accelerate cardiac dysfunction. However, there exists no United States regulatory approved transcatheter device for the treatment AI. The effectiveness of isolated transcatheter mitral therapy in this population is not well understood, thus we aimed to evaluate outcomes for patients with combined AI and MR in comparison to isolated MR that underwent mitral transcatheter edge-to-edge repair (m-TEER). Retrospective data were obtained from Northwell m-TEER registry. A total of 587 patients that underwent m-TEER at four high volume TAVR/TEER centers within the Northwell Health system were included. All patients had severe MR and were divided into two groups: Group 1 with ≥ 3+ AI (AI+) and the Group 2 with <3+ AI (AI-). Echocardiographic outcomes were evaluated at 1 month. Clinical outcomes were evaluated at one month and 1 year. The primary endpoint was death or re-hospitalization at 1 year. 587 patients were included in the study, 92 in the AI+ group. Baseline characteristics were similar in both groups. Approximately two-thirds of patients in the AI+ group demonstrated an improvement in AI severity after isolated mitral therapy. There was no difference in the primary outcome at 1 month or 1 year. There was also no significant difference in NYHA functional class at 1 month between groups. In conclusion, patients that underwent m-TEER with combined MR and AI (AI+) fared well in comparison to isolated mitral valve dysfunction (AI-), with no discernible differences in survival, NYHA class, or re-hospitalization rates at 1 month or 1 year. Hence, isolated m-TEER is a reasonable treatment approach in patients with a high surgical risk with combined AI and MR.

Clinical characteristics and early surgical outcomes of aortoesophageal fistula.

BACKGROUND: The incidence and prognosis of aortoesophageal fistula (AEF) has not been clarified. The clinical characteristics and surgical outcomes of AEF were investigated. METHODS: The clinical data of patients who underwent surgical treatment for AEF from January 2020 to December 2021 that were registered in the Japan Cardiovascular Surgery Database (JCVSD) were analyzed. RESULTS: During the period, 123 patients (71.0 [IQR: 61.0-78.0] years old; 76.4% men) underwent surgical treatment for AEF. The prevalence of secondary AEF was 61%. Secondary AEF after aortic grafting was the most frequent (n = 40; 32.5%), followed by AEF after thoracic endovascular aortic repair (TEVAR) (n = 30; 24.4%). Operative mortality was observed in 23 patients (18.7%). TEVAR for AEF (p = 0.019), postoperative bleeding (p = 0.047), stroke (p = 0.004), renal failure (p < 0.001), newly required hemodialysis (p = 0.023), pneumonia (p = 0.003), multisystem failure (p < 0.001), and dyslipidemia (p = 0.02) were associated with risk factors of operative mortality after surgical treatment of AEF on univariable logistic regression analyses. CONCLUSIONS: This first nationwide study on the surgical treatment for AEF demonstrated a higher incidence of secondary AEF than primary AEF. Both open surgical repair and TEVAR for AEF were associated with high operative mortality. TEVAR and dyslipidemia were risk factors for operative mortality. Precautions and further improved treatment strategies for AEF are still required.

Host repair polymorphisms and H. pylori genes in gastric disease outcomes: Who are the guardian and villains?

Gastric cancer (GC) is the fourth-leading cause of cancer-related mortality. The intestinal subtype of GC comes after the cascade of Correa, presenting H. pylori infection as the major etiological factor. One of the main mechanisms proposed for the progression from a more benign gastric lesion to cancer is DNA damage caused by chronic inflammation. Polymorphisms in DNA repair genes can lead to an imbalance of host DNA damage and repair, contributing to the development of GC. From there, we evaluated the risk of polymorphisms in DNA repair system genes in progressive gastric diseases and their association with the H. pylori genotype. This study included 504 patients from two public hospitals in Brazil's north and northeast regions. The samples were classified into active and inactive gastritis, metaplasia, and GC. Polymorphisms in the DNA repair genes MLH1-93G > A, APE1 2197 T > G, XRCC1 28,152 G > A, MGMT 533 A > G, and XRCC3 18,067C > T were investigated by RFLP-PCR and H. pylori genotype by PCR. Statistical analyses were conducted using EPINFO 7.0., SNPSTAT, and CART software. The XRCC1 (GA) polymorphic allele stood out because it was associated with a lower risk of more severe gastric disease progression. Haplotypes of XRCC1 (GA) associated with some genotypes of MGMT, XRCC3, MLH1, and APE1 also showed protection against the progression of gastric diseases. XRCC3 (CT) showed a decreased risk of gastric disease progression in women, while a risk 1.3x to GC was observed in the MLH1 (A) polymorphic allele. The interaction between H. pylori genes and the host showed that the H. pylori cagE gene was the most important virulence factor associated with a worse clinical outcome, even overlapping with the XRCC1 polymorphism, where the MLH1 polymorphism response varied according to vacA alleles. Our results show the relevance of XRCC1 G > A for genome integrity, sex influence, and interaction between H. pylori virulence factors and XRCC1 and MLH1 genotypes for gastric lesion outcomes in Brazilian populations.

Uncovering the effects of non-lethal oxidative stress on replication initiation in Escherichia coli.

Cell cycle adaptability assists bacteria in response to adverse stress. The effect of oxidative stress on replication initiation in Escherichia coli remains unclear. This work examined the impact of exogenous oxidant and genetic mutation-mediated oxidative stress on replication initiation. We found that 0-0.5 mM H2O2 suppresses E. coli replication initiation in a concentration-dependent manner but does not lead to cell death. Deletion of antioxidant enzymes SodA-SodB, KatE, or AhpC results in delayed replication initiation. The antioxidant N-acetylcysteine (NAC) promotes replication initiation in ΔkatE and ΔsodAΔsodB mutants. We then explored the factors that mediate the inhibition of replication initiation by oxidative stress. MutY, a base excision repair DNA glycosylase, resists inhibition of replication initiation by H2O2. Lon protease deficiency eliminates inhibition of replication initiation mediated by exogenous H2O2 exposure but not by katE or sodA-sodB deletion. The absence of clpP and hslV further delays replication initiation in the ΔktaE mutant, whereas hflK deletion promotes replication initiation in the ΔkatE and ΔsodAΔsodB mutants. In conclusion, non-lethal oxidative stress inhibits replication initiation, and AAA+ proteases are involved and show flexible regulation in E. coli.

AP endonuclease 1: Biological updates and advances in activity analysis.

Apurinic/apyrimidinic endodeoxyribonuclease 1 (APE1, APEX1, REF1, HAP1) is an abasic site-specific endonuclease holding critical roles in numerous biological functions including base excision repair, the DNA damage response, redox regulation of transcription factors, RNA processing, and gene regulation. Pathologically, APE1 expression and function is linked with numerous human diseases including cancer, highlighting the importance of sensitive and quantitative assays to measure APE1 activity. Here, we summarize biochemical and biological roles for APE1 and expand on the discovery of APE1 inhibitors. Finally, we highlight the development of assays to monitor APE1 activity, detailing a recently improved and stabilized DNA Repair Molecular Beacon assay to analyze APE1 activity. The assay is amenable to analysis of purified protein, to measure changes in APE1 activity in cell lysates, to monitor human patient samples for defects in APE1 function, or the cellular and biochemical response to APE1 inhibitors.

Expression, purification, and biochemical analysis of the RNA-DNA hybrid helicase Sen1/SETX from Chaetomium thermophilum.

Yeast Sen1 and its vertebrate ortholog Senataxin (also known as SETX) are RNA-DNA resolving helicases. Sen1 and SETX are implicated in multiple critical nuclear functions not limited to but including DNA replication and repair, RNA processing, and transcription. These> 200 kDa helicases have a two-domain architecture with an N-terminal regulatory helical repeat array linked to an SF1b helicase motor core via a variable sized central linker of low complexity sequence. Given the size of these proteins, production of milligram quantities of protein that is suitable for biochemical, biophysical, and protein structural analysis has been challenging. To overcome these limitations, we developed a robust selectable high-yield YFP-fusion protein expression method for Sen1 production in mammalian cells, followed by purification on a high-affinity YFP-binding camelid nanobody support. Herein, we detail methods and protocols for the expression and purification of recombinant Sen1 from the thermophilic fungus Chaetomium thermophilum, and the quantitative characterization of its RNA-DNA duplex resolution activity.

Utilizing nuclear extracts to characterize protein: DNA interactions at the single molecule level.

Single molecule experiments are invaluable approaches to analyze the dynamics of protein-protein and protein-DNA interactions in real time. SMADNE, single molecule analysis of DNA binding proteins from nuclear extracts, is a new method that allows analysis of a fluorescently tagged overexpressed protein of interest near its native environment while still retaining the advantages of single molecule approaches. Having all the endogenous proteins found in the nucleus provides more biologically relevant information due to their interactions with the protein of interest. Examples of such include the ability for posttranslational modifications to occur, intrinsic stabilization factors, and high labeling efficacy of the protein of interest. Furthermore, having the capabilities to incorporate different DNA substrates and protein variants can elucidate information of the system in a more detailed manner. Finally, orthogonal labeling strategies allows determination of the order of assembly and disassembly of several proteins at sites of damage. This chapter will describe the methodologies, benefits, and applications of SMADNE.

Measuring protein stoichiometry with single-molecule imaging in Xenopus egg extracts.

In human cells, DNA double-strand breaks are rapidly bound by the highly abundant non-homologous end joining (NHEJ) factor Ku70/Ku80 (Ku). Cellular imaging and structural data revealed a single Ku molecule is bound to a free DNA end and yet the mechanism regulating Ku remains unclear. Here, we describe how to utilize the cell-free Xenopus laevis egg extract system in conjunction with single-molecule microscopy to investigate regulation of Ku stoichiometry during non-homologous end joining. Egg extract is an excellent model system to study DNA repair as it contains the soluble proteome including core and accessory NHEJ factors, and efficiently repairs double-strand breaks in an NHEJ-dependent manner. To examine the Ku stoichiometry in the extract system, we developed a single-molecule photobleaching assay, which reports on the number of stable associated Ku molecules by monitoring the intensity of fluorescently labeled Ku molecules bound to double-stranded DNA over time. Photobleaching is distinguishable as step decreases in fluorescence intensity and the number of photobleaching events indicate fluorophore stoichiometry. In this paper we describe sample preparation, experimental methodology, and data analysis to discern Ku stoichiometry and the regulatory mechanism controlling its loading. These approaches can be readily adopted to determine stoichiometry of molecular factors within other macromolecular complexes.

Recovery of insect-pathogenic fungi from solar UV damage: Molecular mechanisms and prospects.

Molecular mechanisms underlying insect-pathogenic fungal tolerance to solar ultraviolet (UV) damage have been increasingly understood. This chapter reviews the methodology established to quantify fungal response to solar UV radiation, which consists of UVB and UVA, and characterize a pattern of the solar UV dose (damage) accumulated from sunrise to sunset on sunny summer days. An emphasis is placed on anti-UV mechanisms of fungal insect pathogens in comparison to those well documented in model yeast. Principles are discussed for properly timing the application of a fungal pesticide to improve pest control during summer months. Fungal UV tolerance depends on either nucleotide excision repair (NER) or photorepair of UV-induced DNA lesions to recover UV-impaired cells in the darkness or the light. NER is a slow process independent of light and depends on a large family of anti-UV radiation (RAD) proteins studied intensively in model yeast but rarely in non-yeast fungi. Photorepair is a rapid process that had long been considered to depend on only one or two photolyases in filamentous fungi. However, recent studies have greatly expanded a genetic/molecular basis for photorepair-dependent photoreactivation that serves as a primary anti-UV mechanism in insect-pathogenic fungi, in which photolyase regulators required for photorepair and multiple RAD homologs have higher or much higher photoreactivation activities than do photolyases. The NER activities of those homologs in dark reactivation cannot recover the severe UV damage recovered by their activities in photoreactivation. Future studies are expected to further expand the genetic/molecular basis of photoreactivation and enrich principles for the recovery of insect-pathogenic fungi from solar UV damage.

Safety and effectiveness of Neuroform Atlas stent-assisted coil embolization for ruptured intracranial aneurysms.

Objective: The treatment outcomes of ruptured intracranial aneurysms using the Neuroform Atlas stent were evaluated. Methods: This study represents a retrospective review that included patients who underwent endovascular treatment for ruptured aneurysms at a single institution. Between January 2018 and September 2022, endovascular treatments including simple coiling or Neuroform Atlas stent-assisted coil embolization were performed in 191 patients with ruptured intracranial aneurysms. Results: Intraprocedural rupture was observed in 11 (8.7%) patients in the Simple Coiling (SC) group, which was slightly higher than that in 4 (6.3%) patients in the Neuroform Atlas stent-assisted coiling (NASAC) group (p=0.241). However, Thromboembolic event (TEE) was slightly more prevalent in the NASAC group, with 4 (6.3%) cases as compared to the 5 (3.9%) cases in the SC group (p=0.235). The retreatment rate was slightly higher in the SC group, with 19 (26.4%) patients as compared to the 10 patients (22.2%) in the NASAC group (p=0.342). Conclusions: The use of the Neuroform Atlas stent (NAS) for ruptured aneurysms might be safe and effective.

[Efficacy analysis of ansa cervicalis anterior root-recurrent laryngeal nerve anastomosis in the treatment of unilateral vocal cord paralysis].

Objective:Investigates the application and clinical efficacy of ansa cervicalis anterior root-recurrent laryngeal nerve（RLN） anastomosis in the treatment of unilateral vocal fold paralysis（UVFP）. Methods:A prospective study was conducted with 92 UVFP patients admitted to our department from January 2018 to January 2022 who received ansa cervicalis anterior root-RLN anastomosis. The course of nerve injury ranged from 6 to 24 months. Videostroboscopy, voice subjective auditory perceptual assessment（GRBAS）, Voice Handicap Index（VHI-10）, voice objective acoustic analysis and laryngeal electromyography（EMG） were used to evaluate the efficacy of the operation. Results:Videostroboscopy showed that although the movement of vocal cords did not return to normal 12 months after operation, their volume and muscle tension were significantly improved and their positions were adducted to the median or near-median. Also the glottic closure, vocal cord position, vocal cord edge, symmetry and regularity of vocal cord vibration were significantly improved than pre-operation（P<0.01）. The five indexes of GRBAS（Grade, Roughness, Breathiness, Asthenia, Strain） and VHI-10, as well as voice acoustic parameters（Jitter, Shimmer, NHR） post-operation were significantly reduced, while the maximum phonation time（MPT） was significantly longer（P<0.01）. The results of laryngeal EMG indicated that the maximum voluntary motor unit recruitment（VMUR） post-operation was significantly recovered（P<0.01）, which confirmed that the affected laryngeal muscle obtained effective nerve reinnervation. Conclusion:Ansa cervicalis anterior root-RLN anastomosis can effectively improve the voice function of patients which is safe and satisfactory. It is an ideal method for the treatment of unilateral RLN injury.

[The current status and development of nerve repair techniques for bilateral vocal cord paralysis].

The related research of nerve repair for bilateral vocal cord paralysis（BVCP） can be traced back to a century ago. There is still no standardized surgical therapeutic schedule and the evaluation of curative effect, due to the variability and complexity of laryngeal nerve innervation. Otolaryngologists have been constantly weighing and improving the surgical plan in relieving airway obstruction, maintaining vocal function and reducing cough to protect the normal physiological function of the larynx. Different medical institutions have great differences in the treatment methods. Many studies and literatures were published on the treatment of BVCP, yet have few related works on the application of it's nerve repair technology presently. The research progress of nerve repair technology for BVCP were described, along with its developing trend, aiming to lay the foundation for the future technical exploration and development.

Surmounting the summit: tackling cerebral malperfusion in AMDS-treated acute deBakey I dissections. A case report.

Background: Acute Type A aortic dissection (ATAAD) with supra-aortic branch (SAB) malperfusion remains a formidable clinical challenge, often resulting in high mortality and complex treatment dilemmas. The introduction of the AMDS represents a significant innovation, designed to stabilize the aortic arch, and manage malperfusion effectively. Methods: This case study evaluates the utility of AMDS in the treatment of a 63-year-old male with hypertension, who presented with severe, acute chest pain. Diagnosed with a DeBakey type I ATAAD involving SAB, the patient underwent cardiopulmonary bypass, aortic root replacement, aortic arch repair with AMDS implantation, and subsequent endovascular stenting for severe left common carotid artery malperfusion that developed postoperatively. The AMDS was instrumental in facilitating crucial aortic arch reconstruction and addressing the initial severe malperfusion. Despite postoperative cerebral malperfusion, targeted endovascular stenting resulted in a rapid and substantial neurological recovery. The patient was discharged to a rehabilitation facility on postoperative day 20, free of neurological deficits. Conclusions: The use of AMDS in managing ATAAD with SAB involvement is transformative, enabling less invasive surgical techniques and offering immediate, effective correction of malperfusion. This case underscores the essential role of integrating advanced endovascular strategies to enhance outcomes in high-risk aortic surgeries, marking a pivotal advancement in the therapeutic approach to complex aortic dissections.

Pre-constrain technique using the Gore Excluder Conformable endograft for abdominal aortic aneurysm with severely angulated proximal neck.

Endovascular aneurysm repair for abdominal aortic aneurysms with a severely angulated proximal neck remains challenging because of the higher rates of type 1a endoleaks and secondary interventions. In this study, we reviewed six consecutive patients with a severely angulated proximal neck (>60°) treated with endovascular aneurysm repair at a single center. They were treated with a Gore Excluder Conformable endograft using a pre-constrain technique to prevent type 1a endoleaks. It is the unique deployment technique of the proximal trunk by pre-constrain of the proximal edge. This technique helps obtain optimal deployment of the proximal main body.

Long head of biceps tendon augmentation in rotator cuff repair enhances tendon healing, shoulder function and patient-reported outcomes one-year post-surgery.

Purpose: The aim is to determine the effect on healing and functionality of patients after 1 year of biceps augmentation of a rotator cuff repair (RCR) compared to RCR plus long head of the biceps (LHB) tenotomy. In addition, to analyse the main factors involved in the recovery after the surgery. Methods: A prospective, comparative, non-randomized study (Level of Evidence III) was conducted. Patients with repairable rotator cuff tears were allocated to either the control group, with a double row transosseous equivalent RCR with LHB tenotomy, or the RCR+augmentation with LHB group. Patients were evaluated for radiological (MRI), clinical (cuff size, Patte and Goutallier scales) and functional variables (Constant and American Shoulder and Elbow Surgeons [ASES] scales) before the intervention. At 1-year follow-up cuff healing was confirmed through MRI and functional evaluation with Constant, ASES, simple shoulder test [SST] and Disabilities of the Arm, Shoulder and Hand scales. Results: Seventy-seven patients underwent control or RCR+augmentation with LHB, there were no preoperative differences between the groups. After 1 year of the surgery, re-rupture occurred in 38.5% and 16% of the patients in control and RCR+augmentation with LHB groups, respectively (p = .026). Total functionality was higher (p < .05) in RCR+augmentation with LHB than in the control group: Constant, SST and ASES scales. Among the explored factors involved in healing, re-rupture occurred in 100% of the cases with high fatty degeneration. Besides, higher initial functionality (Constant scale) and RCR+augmentation with LHB increased the odds of healing (odds ratio [OR] = 1.12 [1.04-1.21]; OR = 5 [1, 61], respectively), while higher cuff length had a detrimental effect (OR = 0.92 [0.85-0.99]). Conclusion: RCR+augmentation with LHB achieves a higher healing percentage and a better functional evolution than RCR+LHB tenotomy, 1 year after cuff repair. Fatty degeneration, cuff length and initial functionality are the main factors involved in cuff healing. Level of Evidence: Level III randomized controlled trial.