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In the field of sustainable hydrogen production, hematite (α-Fe2O3) is a promising material owing to its optical band gap and water oxidation/reduction energies. In this context, the main objective of this work is devoted to the synthesis of α-Fe2O3 by electrochemical anodization of iron sheet using two anodization voltages (20 and 30 V), followed by annealing at 450°C (1.5 h) in air. Indeed, annealing is a necessary but not sufficient condition for the hematite prepared electrochemically. Furthermore, we discuss the effect of anodizing voltage and analyze its influence on the properties of the films namely structural, morphological, optical, electrochemical and photoelectrochemical (PEC) properties. The annealed Fe2O3 sample, anodized at 30 V, demonstrates a net photocurrent density of 0.81 mA/cm2 at 0.744 VRHE in KOH (1 M). The obtained results indicate that the α-hematite thin films synthesized by anodization technique under optimized conditions is a promising photoanode in PEC cells. [ABSTRACT FROM AUTHOR]

Cognitive impairment is one of the main complications of diabetes and patients with cognitive impairment are considered to be in an intermediate stage of decreased cognition with higher risk of developing dementia and Alzheimer's disease. This study aimed at evaluating serum activity of two esterase enzymes including cholinesterase and paraoxonase 1 (PON 1) in diabetic patients and association with cognitive performance and metabolic parameters. In this crosssectional study, 128 diabetic patients were enrolled. Cognitive performance was evaluated using the Wechsler Adult Intelligence Scale-Revised (WAIS-R). Body mass index, fasting blood glucose, fasting blood insulin, insulin resistance, serum cholinesterase and paraoxonase 1 activity were measured. Prevalence of cognitive impairment was found to be 77% in the study population. Serum cholinesterase activity was found to be higher in diabetic patients with cognitive impairment than the patients with normal cognitive performance (p < 0.04). Further, a negative correlation was found between cognitive performance and cholinesterase activity, albeit statistically non-significant. Alteration in the activity of cholinesterase enzyme was shown to be associated with cognitive impairment in diabetic patients and future studies are required to find the pathophysiological mechanisms of diabetic cognitive dysfunction. [ABSTRACT FROM AUTHOR]

Synthesis of mechanically stable porous scaffolds with an architecture analogous to cancellous bone tissue poses significant challenges to bioactive glass (BG) based scaffolds. This is primarily due to densification and crystallization of the BG's during heat treatment. This study presents a modified BG series (42SiO 2 -xTiO 2 -24Na 2 O-21CaO-13P 2 O 5 , where x = 8 and 16 TiO 2 ). TiO 2 replaced the SiO 2 concentration in the glass and was incorporated due to its biocompatibility and influence on glass structure. Material characterization determined that TiO 2 did not induce crystallization within the glass but did increase the glass transition temperature (T g ) from 520°C to 600°C thereby indicating a more stable network connectivity. Scaffolds were synthesized using the foam replication method, resulting in scaffolds with a pore size of approximately 500 μm with the BG-4 composition (30SiO 2 -12TiO 2 -24Na 2 O-21CaO-13P 2 O 5 ) retaining its amorphous character post-heat treatment. Scaffold ion release was monitored over 5-60 days in simulated body fluid (SBF). Si 4+ release was found to decrease, while Ca 2+ levels increased in SBF as TiO 2 replaced SiO 2 within the glass series. Cytocompatibility studies revealed that MC3T3 Osteoblast cells proliferated on the BG-4 scaffold surface and at its interface within culture media, and cell numbers were not significantly reduced., (© 2024 Wiley Periodicals LLC.)

The optimal myeloablative conditioning regimen for ALL patients undergoing hematopoietic cell transplant (HCT) with an alternative donor is unknown. We analyzed HCT outcomes ALL patients (n = 269) who underwent HCT at our center from 2010 to 2020 in complete remission (CR) after FTBI-etoposide and CNI-based GvHD prophylaxis for matched donor HCT (ETOP-package; n = 196) or FTBI-Fludarabine and post-transplant cyclophosphamide (PTCy)-based prophylaxis for HLA- mismatched (related or unrelated) donors (FLU-package; n = 64). Patients in FLU-package showed a significant delay in engraftment (p < 0.001) and lower cumulative incidence (CI) of any and extensive chronic GVHD (p = 0.009 and 0.001, respectively). At the median follow up of 4.6 years (range 1-12 years); non-relapse mortality, overall or leukemia-free survival and GVHD-free/relapse-free survival were not significantly impacted by the choice of conditioning. However, in patients at CR2 or with measurable residual disease (MRD+), there was a trend towards higher relapse after FLU-package (p = 0.08 and p = 0.07, respectively), while patients at CR1 regardless of MRD status had similar outcomes despite the package/donor type (p = 0.9 and 0.7, respectively). Our data suggests that FLU-package for alternative donors offers comparable outcomes to ETOP-package for matched donor HCT to treat ALL. Disease status and depth of remission at HCT were independent predictors for better outcomes., (© 2024. The Author(s).)

Allogeneic hematopoietic cell transplantation (HCT) remains the sole curative treatment for most patients with hematologic malignancies. A well-matched donor (related or unrelated) remains the preferred donor for patients undergoing allogeneic HCT; however, a large number of patients rely on alternative donor choices of mismatched related (haploidentical) or unrelated donors to access HCT. In this retrospective study, we investigated outcomes of patients who underwent mismatched donor (related or unrelated) HCT with a radiation-based myeloablative conditioning MAC regimen in combination with fludarabine, and post-transplantation cyclophosphamide (PTCy) as higher-intensity graft-versus-host disease (GVHD) prophylaxis. We retrospectively assessed HCT outcomes in 155 patients who underwent mismatched donor HCT (related/haploidentical versus unrelated [MMUD]) with fractionated-total body irradiation (fTBI) plus fludarabine and PTCy as GVHD prophylaxis at City of Hope from 2015 to 2021. Diagnoses included acute lymphoblastic leukemia (46.5%), acute myelogenous leukemia (36.1%), and myelodysplastic syndrome (6.5%). The median age at HCT was 38 years, and 126 patients (81.3%) were an ethnic minority. The Hematopoietic Stem Cell Transplantation Comorbidity Index was ≥3 in 36.1% of the patients, and 29% had a Disease Risk Index (DRI) of high/very high. The donor type was haploidentical in 67.1% of cases and MMUD in 32.9%. At 2 years post-HCT, disease-free survival (DFS) was 75.4% and overall survival (OS) was 80.6% for all subjects. Donor type did not impact OS (hazard ratio [HR], .72; 95% confidence interval [CI], .35 to 1.49; P = .37) and DFS (HR, .78; 95% CI, .41 to 1.48; P = .44), but younger donors was associated with less grade III-IV acute GVHD (HR, 6.60; 95% CI, 1.80 to 24.19; P = .004) and less moderate or severe chronic GVHD (HR, 3.53; 95% CI, 1.70 to 7.34; P < .001), with a trend toward better survival (P = .099). The use of an MMUD was associated with significantly faster neutrophil recovery (median, 15 days versus 16 days; P = .014) and platelet recovery (median, 18 days versus 24 days; P = .029); however, there was no difference in GVHD outcomes between the haploidentical donor and MMUD groups. Nonrelapse mortality (HR, .86; 95% CI, .34 to 2.20; P = .76) and relapse risk (HR, .78; 95% CI, .33 to 1.85; P = .57) were comparable in the 2 groups. Patient age <40 years and low-intermediate DRI showed a DFS benefit (P = .004 and .029, respectively). High or very high DRI was the only predictor of increased relapse (HR, 2.89; 95% CI, 1.32 to 6.34; P = .008). In conclusion, fludarabine/fTBI with PTCy was well-tolerated in mismatched donor HCT, regardless of donor relationship to the patient, provided promising results, and increased access to HCT for patients without a matched donor, especially patients from ethnic minorities and patients of mixed race., (Copyright © 2024 The American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. All rights reserved.)

Purpose: To assess the fracture resistance and failure mode of endodontically treated premolars reconstructed by different preparation approaches: cervical margin relocation (CMR) and crown lengthening (CL) with complete ferrule (CLF) and partial ferrule (CLPF) with three different post and core systems., Materials and Methods: In this in vitro study, 100 maxillary premolars were assigned to the following 10 groups according to their preparation approach and type of post and core system (n = 10): (I) control (intact teeth), (II) prefabricated fiber post (PFP) and composite core with CMR (PFP-CMR), (III) polyethylene fiber-reinforced composite (PEFRC) with CMR (PEFRC-CMR), (IV) casting post (CP) and core with CMR (CP-CMR), (V) PFP-CLPF, (VI) PEFRC-CLPF, (VII) CP-CLPF, (VIII) PFP-CLF, (IX) PEFRC-CLF, and (X) CP-CLF. After thermomechanical loading, the fracture resistance and failure mode were assessed. Data were analyzed statistically (α = 0.05)., Results: In all post and core systems, the CLPF approach had lower fracture resistance than CMR (p < 0.05); CLF showed higher fracture resistance than CLPF only in the PFP system (p = 0.038). In PEFRC and CP systems, the difference between CLF and CLPF was not significant (p > 0.05). No significant difference was found in fracture resistance of different post and core systems with the same preparation approach (p > 0.05). CLPF showed the highest frequency of favorable, and CLF showed the highest frequency of unfavorable fractures., Conclusion: CLPF yielded lower fracture resistance than CMR. The difference in fracture resistance was not significant between CLF and CMR but the frequency of unfavorable fractures was higher in CLF than in other groups., (© 2023 by the American College of Prosthodontists.)

Background: Mesenchymal stromal cells (MSCs) exert immunomodulatory effects, primarily through released extracellular vesicles (EVs). For the clinical-grade manufacturing of MSC-EV products culture conditions need to support MSC expansion and allow the manufacturing of potent MSC-EV products. Traditionally, MSCs are expanded in fetal bovine serum-supplemented media. However, according to good manufacturing practice (GMP) guidelines the use of animal sera should be avoided. To this end, human platelet lysate (hPL) has been qualified as an animal serum replacement. Although hPL outcompetes animal sera in promoting MSC expansion, hPL typically contains components of the coagulation system that need to be inhibited or removed to avoid coagulation reactions in the cell culture. Commonly, heparin is utilized as an anticoagulant; however, higher concentrations of heparin can negatively impact MSC viability, and conventional concentrations alone do not sufficiently prevent clot formation in prepared media., Methods: To circumvent unwanted coagulation processes, this study compared various clotting prevention strategies, including different anticoagulants and calcium chloride (CaCl 2 )-mediated declotting methods, which in combination with heparin addition was found effective. We evaluated the influence of the differently treated hPLs on the proliferation and phenotype of primary bone marrow-derived MSCs and identified the CaCl 2 -mediated declotting method as the most effective option. To determine whether CaCl 2 declotted hPL allows the manufacturing of immunomodulatory MSC-EV products, EVs were prepared from conditioned media of MSCs expanded with either conventional or CaCl 2 declotted hPL. In addition to metric analyses, the immunomodulatory potential of resulting MSC-EV products was assessed in a recently established multi-donor mixed lymphocyte reaction assay., Results and Conclusions: Our findings conclusively show that CaCl 2 -declotted hPLs support the production of immunomodulatory-active MSC-EV products., Competing Interests: Declaration of Competing Interest BG is a scientific advisory board member of Innovex Therapeutics SL, Mursla Ltd, ReNeuron Ltd and PL BioScience. He is a founding director of Exosla Ltd. HH is founder and executive director at PL Bioscience a company, selling hPLs as cell culture supplements. All other authors report no conflicts of interest., (Copyright © 2024 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.)

Research frequently uses words as stimuli to assess cognitive and psychological processes. However, various attributes of these words, such as their semantic and emotional aspects, could potentially confound study results if not properly controlled. This study aims to establish a reliable foundation for the semantic and emotional aspects of words for research in Persian. To this end, the present study provided norms for 718 Persian nouns in arousal, valence, familiarity, and animacy dimensions. The words were selected from a previous English dataset (Warriner et al. in Behav Res Methods 45(4):1191-1207, 2013), translated into Persian, and rated by a total of 463 native Persian-speaking participants. The ratings were obtained through an online questionnaire using a 9-point Likert scale for emotional dimensions (i.e., valence and arousal) and a 5-point Likert scale for semantic dimensions (i.e., familiarity and animacy). The reliability of the ratings was measured using the split-half method, and the result indicated a high consistency of ratings in all dimensions. To assess the relationship between the emotional and semantic dimensions, Pearson correlation coefficient was conducted. Gender differences were investigated through the Mann-Whitney U test, and significant differences were observed in all dimensions. These results are compared with findings from previous studies that were conducted in various languages., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Objective: This study aimed to assess the effect of length of the relocated cervical margin with casting post and core (CP), prefabricated fiber post and composite core (PFP), and polyethylene fiber-reinforced composite (PEFRC) on fracture resistance and marginal integrity., Materials and Methods: In this in vitro study, 70 sound human maxillary premolars were divided into seven groups according to the type of post and core system and length of the relocated cervical margin ( n  = 10): control (no preparation), PFP-3, PEFRC-3, CP-3 with 3 mm of cervical margin relocation (CMR), PFP-6, PEFRC-6, and CP-6 (with 6 mm of CMR). The samples were restored with zirconia crowns (except the control group). Epoxy resin replicas were fabricated before and after thermomechanical loading. Marginal integrity was assessed at the luting cement-core, core-tooth, and luting cement-enamel interfaces under a scanning electron microscope (SEM) (×200). Fracture resistance and failure mode were subsequently assessed. Data were analyzed by independent t -test, paired t -test, ANOVA, Tukey-Games Howell, Mann-Whitney, Kruskal-Wallis, Wilcoxon, Mann-Whitney with Bonferroni correction, and Fisher-Freeman-Halton tests ( α  = 0.05)., Results: The marginal integrity of the groups with 3 mm of CMR followed the following order: PEFRC > PFP > CP at all interfaces ( P   < 0.05). In 6-mm CMR groups, this order was CP < PFP = PEFRC at the luting cement-core and (CP < PEFRC) = PFP at the core-tooth interface. No significant difference was found in fracture resistance when comparing the 3-mm CMR groups with each other ( P   > 0.05). PFP-6 showed higher FR than CP-6 ( P   < 0.001). PEFRC-6 had no significant difference with PFP-6 and CP-6 ( P   > 0.05). In each post and core system, 3-mm CMR groups showed higher marginal integrity and fracture resistance ( P   < 0.05)., Conclusion: Increasing the length of the relocated cervical margin decreased the marginal integrity and fracture resistance of all three systems of CP, PFP, and PEFRC., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2024 Naghmeh Musapoor et al.)

Advanced glycation end products (AGEs) are the final products of the Maillard reaction, formed through the interaction of carbohydrates and proteins. Reactive dicarbonyl compounds such as methylglyoxal (MGO) serve as precursors for AGEs formation. Elevated levels of MGO/AGEs are observed in conditions like obesity, polycystic ovarian syndrome (PCOS), and diabetes, negatively impacting oocyte development. Previous studies have shown that hydrogen sulfide, a gasotransmitter with anti-AGEs effects, is produced in a process influenced by vitamin B6. R-α-lipoic acid (ALA) inhibits protein glycation and AGEs formation while stimulating glutathione (GSH) production. Taurine mitigates oxidative stress and acts as an anti-glycation compound, preventing in vitro glycation and AGEs accumulation. This study aimed to explore the ameliorative effects of a micronutrient support (Taurine, ALA and B6: TAB) on mouse oocytes challenged with MGO. Our results indicate that MGO reduces oocyte developmental competence, while TAB supplementation improves maturation, fertilization, and blastocyst formation rates. TAB also restores cell lineage allocation, redox balance and mitigates mitochondrial dysfunction in MGO-challenged oocytes. Furthermore, cumulus cells express key enzymes in the transsulfuration pathway, and TAB enhances their mRNA expression. However, TAB does not rescue MGO-induced damage in denuded oocytes, emphasizing the supportive role of cumulus cells. Overall, these findings suggest that TAB interventions may have significant implications for addressing reproductive dysfunctions associated with elevated MGO/AGEs levels. This study highlights the potential of TAB supplementation in preserving the developmental competence of COCs exposed to MGO stress, providing insights into mitigating the impact of dicarbonyl stress on oocyte quality and reproductive outcomes., (© 2024. The Author(s).)

Wild-type Lactococcus lactis strain LAC460 secretes prophage-encoded bacteriocin-like lysin LysL, which kills some Lactococcus strains, but has no lytic effect on the producer. LysL carries two N-terminal enzymatic active domains (EAD), and an unknown C-terminus without homology to known domains. This study aimed to determine whether the C-terminus of LysL carries a cell wall binding domain (CBD) for target specificity of LysL. The C-terminal putative CBD region of LysL was fused with His-tagged green fluorescent protein (HGFPuv). The HGFPuv&#95;CBDlysL gene fusion was ligated into the pASG-IBA4 vector, and introduced into Escherichia coli. The fusion protein was produced and purified with affinity chromatography. To analyse the binding of HGFPuv&#95;CBDLysL to Lactococcus cells, the protein was mixed with LysL-sensitive and LysL-resistant strains, including the LysL-producer LAC460, and the fluorescence of the cells was analysed. As seen in fluorescence microscope, HGFPuv&#95;CBDLysL decorated the cell surface of LysL-sensitive L. cremoris MG1614 with green fluorescence, whereas the resistant L. lactis strains LM0230 and LAC460 remained unfluorescent. The fluorescence plate reader confirmed the microscopy results detecting fluorescence only from four tested LysL-sensitive strains but not from 11 tested LysL-resistant strains. Specific binding of HGFPuv&#95;CBDLysL onto the LysL-sensitive cells but not onto the LysL-resistant strains indicates that the C-terminus of LysL contains specific CBD. In conclusion, this report presents experimental evidence of the presence of a CBD in a lactococcal phage lysin. Moreover, the inability of HGFPuv&#95;CBDLysL to bind to the LysL producer LAC460 may partly explain the host's resistance to its own prophage lysin., (© 2024. The Author(s).)

Lung cancer is a leading cause of cancer-related morbidity and mortality worldwide. Metastases in the brain are a common hallmark of advanced stages of the disease, contributing to a dismal prognosis. Lung cancer can be broadly classified as either small cell lung cancer (SCLC) or non-small cell lung cancer (NSCLC). NSCLC represents the most predominant histology subtype of lung cancer, accounting for the majority of lung cancer cases. Recent advances in molecular genetics, coupled with innovations in small molecule drug discovery strategies, have facilitated both the molecular classification and precision targeting of NSCLC based on oncogenic driver mutations. Furthermore, these precision-based strategies have demonstrable efficacy across the blood-brain barrier, leading to positive outcomes in patients with brain metastases. This review provides an overview of the clinical features of lung cancer brain metastases, as well as the molecular mechanisms that drive NSCLC oncogenesis. We also explore how precision medicine-based strategies can be leveraged to improve NSCLC brain metastases.

Introduction: Obsessive-compulsive disorder (OCD) is a disabling mental condition. Many studies have shown that OCD patients have cognitive deficits in various aspects of their cognition, which is a worsening factor of symptom severity, a potential endophenotype, insight predictor, and prognostic indicator of OCD. We designed this systematic review to evaluate the clinical efficacy of cognitive rehabilitation in cognitive deficits and symptom severity of patients with OCD following the PRISMA guidelines., Methods: We searched PubMed, Scopus, ScienceDirect, Google Scholar, and Cochrane Library using the MeSH terms and keywords of 'cognitive rehabilitation' and 'obsessive-compulsive disorder'. The database search identified 200 records of interest, and then 105 duplicates were removed from them. From 95 remaining studies, six articles were eligible for the study and met the inclusion criteria. The six articles described individual RCT studies representing a wide variety of study designs., Results: The six included studies investigated the effect of "organizational training", "cognitive remediation", "attention splitting" and "goal management training" on cognitive impairments and symptom severity of OCD patients.There are a small number of studies with different designs and some biases that have examined the effectiveness of cognitive rehabilitation in OCD patients, with conflicting results regarding the effect of cognitive rehabilitation on OCD symptom severity or cognitive deficit., Conclusion: According to the results, we cannot conclude about the efficacy of cognitive rehabilitation in adults with OCD. Considering the importance of cognitive deficits in OCD patients, it is necessary to design and conduct standard trials to investigate the role of cognitive rehabilitation in these disorders., (Copyright© 2024 Iranian Neuroscience Society.)

Background: Respiratory distress syndrome (RDS) is one of the most important and common disorders among premature infants., Objective: This study aimed to compare the effect of the combination of surfactant and budesonide with surfactant alone on Bronchopulmonary dysplasia (BPD) and mortality rate among premature infants with RDS., Method: An outcome assessor-blind randomized clinical trial was conducted on 134 premature infants with RDS who were born in Ayatollah Mousavi Hospital, Zanjan, Iran in 2021. The covariate adaptive randomization method was utilized to allocate participants into two groups (surfactant alone and a combination of surfactant and budesonide). The primary outcomes were BPD and Mortality rate from admission to hospital discharge. The data in this study were analyzed using SPSS software version 18., Results: Overall the comparison of mortality rate and BPD between the two groups did not show a significant difference(p > 0.05). The subgroup results showed that administering surfactant with budesonide to infants under 30 weeks of age significantly reduced the number of deaths compared to using surfactant alone (5 vs. 17). Similar positive effects were observed for the occurrence of Pulmonary Hemorrhage, the need for a second dose of surfactant, oxygen index, mean blood pressure and mean arterial pressure (MAP) in infants under 34 weeks of age compared to more than 34 weeks (p < 0.05)., Conclusion: These findings suggest that the combination therapy of surfactant and budesonide may be beneficial, particularly in preterm infants with less than 34 weeks gestational age and 1500 birth weight. However, further studies with larger sample sizes and longer follow-up periods are needed to confirm these results and assess long-term outcomes., Trial Registration: The study was registered at the Iranian Registry of Clinical Trials website under the code IRCT20201222049802N1. https://en.irct.ir/user/trial/48117/view ., Registration Date: 28/02/2021., Public Repository: DATA SET: This research data set link is displayed on the Zanjan-Iran Medical Sciences website: https://repository.zums.ac.ir/cgi/users/login? target=https%3 A%2 F/repository.zums.ac.ir/id/eprint ., (© 2024. The Author(s).)

Background: Hospitals should have effective and efficient organizational charts to face the changing healthcare environment. Thus, for this purpose, the present study seeks to compile an organizational chart for Iranian hospitals., Materials and Methods: The present study was conducted in two phase overview and qualitative (using focus group discussion). In the overview phase, the organizational charts of hospitals were analyzed in terms of complexity (i.e., degree of horizontal and vertical separations), and the initial hospital organizational chart was developed based on the results. Subsequently, experts were interviewed in a focus group discussion to finalize and validate the initial organizational chart., Results: The final organizational chart was designed to contain features such as internal divisions, specialization, reduction of organizational hierarchies, expansion of supervision scope, and moderate-sized organizational pyramid., Conclusion: Using designed organizational chart would eliminate the redundant managerial levels since it reduces organizational hierarchies to two levels of management, expands the supervision scopes, fosters a moderate-sized organizational pyramid, and catalyzes communications., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Amini-Rarani et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Paraneoplastic neurological syndromes (PNS) are defined as remote neurologic immune-mediated effects triggered by underlying systemic tumors. While recognizing specific syndromes can aid early cancer detection, overutilization of paraneoplastic assays in the absence of a classic syndrome can precipitate overdiagnosis and overtreatment. PNS involve autoantibodies targeting intracellular or extracellular antigens, with variable immunotherapy responses based on antigen type. Diagnosing PNS is challenging, requiring exclusion of other differential diagnoses. New diagnostic criteria classify PNS into high-risk and intermediate-risk phenotypes based on clinical phenotype, neuronal antibodies, and cancer presence. Patients with cell surface antibodies respond better to immunotherapies compared to those with intracellular antigen targets. Understanding PNS syndromes, serological markers, and oncological features guides management, which facilitates initiation of immunosuppression for PNS alongside treatment of the underlying neoplasm, thereby improving neurologic and oncologic outcomes. Initial treatments often include intravenous methylprednisolone, plasma exchange, or intravenous immunoglobulins. Second-line immunosuppressants like rituximab or cyclophosphamide may be necessary if initial treatments fail. Specific therapies vary based on antibody target. Here, we summarize the current approach to the investigation, diagnosis, and treatment of patients with suspected PNS., Competing Interests: None declared., (Thieme. All rights reserved.)

The wild-type Lactococcus lactis strain LAC460 produces two bacteriocin-like phage lysins, LysL and LysP. This study aimed to produce and secrete LysL in various heterologous hosts and an in vitro cell-free expression system for further functional studies. Initially, the lysL gene from L. lactis LAC460 was cloned into Lactococcus cremoris NZ9000 and L. lactis N8 strains, with and without the usp45 signal sequence (SSusp45), under a nisin-inducible promoter. Active LysL was primarily produced intracellularly in recombinant L. lactis N8, with some secretion into the supernatant. Recombinant L. cremoris NZ9000 lysed upon nisin induction, indicating successful lysL expression. However, fusion with Usp45 signal peptide (SPUsp45-LysL) weakened LysL activity, likely due to incomplete signal peptide cleavage during secretion. Active LysL was also produced in vitro, and analysed in SDS-PAGE, giving a 42-kDa band. However, the yield of LysL protein was still low when produced from recombinant lactococci or by in vitro expression system. Therefore, His-tagged LysL was produced in Escherichia coli BL21(DE3). Western blot confirmed the intracellular production of about 44-kDa His-tagged LysL in E. coli. His-tagged active LysL was then purified by Ni-NTA affinity chromatography yielding sufficient 4.34 mg of protein to be used in future functional studies., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)