bluuebunny/crossref_metadata_2025 huggingface dataset:
High-throughput screening of catalytically active inclusion bodies using laboratory automation and Bayesian optimization
Authors: Helleckes Laura Marie , Küsters Kira , Wagner Christian , Hamel Rebecca , Saborowski Ronja , Marienhagen Jan , Wiechert Wolfgang , Oldiges Marco
Published: February 2024

Background

In recent years, the production of inclusion bodies that retain substantial catalytic activity was demonstrated. These catalytically active inclusion bodies (CatIBs) are formed by genetic fusion of an aggregation-inducing tag to a gene of interest via short linker polypeptides. The resulting CatIBs are known for their easy and cost-efficient production, recyclability as well as their improved stability. Recent studies have outlined the cooperative effects of linker and aggregation-inducing tag on CatIB activities. However, no a priori prediction is possible so far to indicate the best combination thereof. Consequently, extensive screening is required to find the best performing CatIB variant.

Results

In this work, a semi-automated cloning workflow was implemented and used for fast generation of 63 CatIB variants with glucose dehydrogenase of Bacillus subtilis (BsGDH). Furthermore, the variant BsGDH-PT-CBDCell was used to develop, optimize and validate an automated CatIB screening workflow, enhancing the analysis of many CatIB candidates in parallel. Compared to previous studies with CatIBs, important optimization steps include the exclusion of plate position effects in the BioLector by changing the cultivation temperature. For the overall workflow including strain construction, the manual workload could be reduced from 59 to 7 h for 48 variants (88%). After demonstration of high reproducibility with 1.9% relative standard deviation across 42 biological replicates, the workflow was performed in combination with a Bayesian process model and Thompson sampling. While the process model is crucial to derive key performance indicators of CatIBs, Thompson sampling serves as a strategy to balance exploitation and exploration in screening procedures. Our methodology allowed analysis of 63 BsGDH-CatIB variants within only three batch experiments. Because of the high likelihood of TDoT-PT-BsGDH being the best CatIB performer, it was selected in 50 biological replicates during the three screening rounds, much more than other, low-performing variants.

Conclusions

At the current state of knowledge, every new enzyme requires screening for different linker/aggregation-inducing tag combinations. For this purpose, the presented CatIB toolbox facilitates fast and simplified construction and screening procedures. The methodology thus assists in finding the best CatIB producer from large libraries in short time, rendering possible automated Design-Build-Test-Learn cycles to generate structure/function learnings.
Recombinant L. lactis vaccine LL-plSAM-WAE targeting four virulence factors provides mucosal immunity against H. pylori infection
Authors: Zhang Furui , Ni Linhan , Zhang Zhen , Luo Xuegang , Wang Xuequan , Zhou Wenmiao , Chen Jiale , Liu Jing , Qu Yuliang , Liu Kunmei , Guo Le
Published: February 2024

Background

Helicobacter pylori (H. pylori) causes chronic gastric disease. An efficient oral vaccine would be mucosa-targeted and offer defense against colonization of invasive infection in the digestive system. Proteolytic enzymes and acidic environment in the gastrointestinal tract (GT) can, however, reduce the effectiveness of oral vaccinations. For the creation of an edible vaccine, L. lactis has been proposed as a means of delivering vaccine antigens.

Results

We developed a plSAM (pNZ8148-SAM) that expresses a multiepitope vaccine antigen SAM-WAE containing Urease, HpaA, HSP60, and NAP extracellularly (named LL-plSAM-WAE) to increase the efficacy of oral vaccinations. We then investigated the immunogenicity of LL-plSAM-WAE in Balb/c mice. Mice that received LL-plSAM-WAE or SAM-WAE with adjuvant showed increased levels of antibodies against H. pylori, including IgG and sIgA, and resulted in significant reductions in H. pylori colonization. Furthermore, we show that SAM-WAE and LL-plSAM-WAE improved the capacity to target the vaccine to M cells.

Conclusions

These findings suggest that recombinant L. lactis could be a promising oral mucosa vaccination for preventing H. pylori infection.
Evaluating oleaginous yeasts for enhanced microbial lipid production using sweetwater as a sustainable feedstock
Authors: Keita Valériane Malika , Lee Yi Qing , Lakshmanan Meiyappan , Ow Dave Siak-Wei , Staniland Paul , Staniland Jessica , Savill Ian , Tee Kang Lan , Wong Tuck Seng , Lee Dong-Yup
Published: February 2024

Background

Yeasts exhibit promising potential for the microbial conversion of crude glycerol, owing to their versatility in delivering a wide range of value-added products, particularly lipids. Sweetwater, a methanol-free by-product of the fat splitting process, has emerged as a promising alternative feedstock for the microbial utilization of crude glycerol. To further optimize sweetwater utilization, we compared the growth and lipid production capabilities of 21 oleaginous yeast strains under different conditions with various glycerol concentrations, sweetwater types and pH.

Results

We found that nutrient limitation and the unique carbon composition of sweetwater boosted significant lipid accumulation in several strains, in particular Rhodosporidium toruloides NRRL Y-6987. Subsequently, to decipher the underlying mechanism, the transcriptomic changes of R. toruloides NRRL Y-6987 were further analyzed, indicating potential sugars and oligopeptides in sweetwater supporting growth and lipid accumulation as well as exogenous fatty acid uptake leading to the enhanced lipid accumulation.

Conclusion

Our comparative study successfully demonstrated sweetwater as a cost-effective feedstock while identifying R. toluroides NRRL Y-6987 as a highly promising microbial oil producer. Furthermore, we also suggested potential sweetwater type and strain engineering targets that could potentially enhance microbial lipid production.

Graphical abstract
A genome-reduced Corynebacterium glutamicum derivative discloses a hidden pathway relevant for 1,2-propanediol production
Authors: Siebert Daniel , Glawischnig Erich , Wirth Marie-Theres , Vannahme Mieke , Salazar-Quirós Álvaro , Weiske Annette , Saydam Ezgi , Möggenried Dominik , Wendisch Volker F. , Blombach Bastian
Published: February 2024

Background

1,2-propanediol (1,2-PDO) is widely used in the cosmetic, food, and drug industries with a worldwide consumption of over 1.5 million metric tons per year. Although efforts have been made to engineer microbial hosts such as Corynebacterium glutamicum to produce 1,2-PDO from renewable resources, the performance of such strains is still improvable to be competitive with existing petrochemical production routes.

Results

In this study, we enabled 1,2-PDO production in the genome-reduced strain C. glutamicum PC2 by introducing previously described modifications. The resulting strain showed reduced product formation but secreted 50 ± 1 mM d-lactate as byproduct. C. glutamicum PC2 lacks the d-lactate dehydrogenase which pointed to a yet unknown pathway relevant for 1,2-PDO production. Further analysis indicated that in C. glutamicum methylglyoxal, the precursor for 1,2-PDO synthesis, is detoxified with the antioxidant native mycothiol (MSH) by a glyoxalase-like system to lactoylmycothiol and converted to d-lactate which is rerouted into the central carbon metabolism at the level of pyruvate. Metabolomics of cell extracts of the empty vector-carrying wildtype, a 1,2-PDO producer and its derivative with inactive d-lactate dehydrogenase identified major mass peaks characteristic for lactoylmycothiol and its precursors MSH and glucosaminyl-myo-inositol, whereas the respective mass peaks were absent in a production strain with inactivated MSH synthesis. Deletion of mshA, encoding MSH synthase, in the 1,2-PDO producing strain C. glutamicum ΔhdpAΔldh(pEKEx3-mgsA-yqhD-gldA) improved the product yield by 56% to 0.53 ± 0.01 mM1,2−PDO mMglucose−1 which is the highest value for C. glutamicum reported so far.

Conclusions

Genome reduced-strains are a useful basis to unravel metabolic constraints for strain engineering and disclosed in this study the pathway to detoxify methylglyoxal which represents a precursor for 1,2-PDO production. Subsequent inactivation of the competing pathway significantly improved the 1,2-PDO yield.
Unlocking Nature’s Toolbox: glutamate-inducible recombinant protein production from the Komagatella phaffii PEPCK promoter
Authors: Rajak Neetu , Dey Trishna , Sharma Yash , Bellad Vedanth , Rangarajan Pundi N.
Published: February 2024

Background

Komagataella phaffii (a.k.a. Pichia pastoris) harbors a glutamate utilization pathway in which synthesis of glutamate dehydrogenase 2 and phosphoenolpyruvate carboxykinase (PEPCK) is induced by glutamate. Glutamate-inducible synthesis of these enzymes is regulated by Rtg1p, a cytosolic, basic helix-loop-helix protein. Here, we report food-grade monosodium glutamate (MSG)-inducible recombinant protein production from K. phaffii PEPCK promoter (PPEPCK) using green fluorescent protein (GFP) and receptor binding domain of SARS-CoV-2 virus (RBD) as model proteins.

Results

PPEPCK-RBD/GFP expression cassette was integrated at two different sites in the genome to improve recombinant protein yield from PPEPCK. The traditional, methanol-inducible alcohol oxidase 1 promoter (PAOX1) was used as the benchmark. Initial studies carried out with MSG as the inducer resulted in low recombinant protein yield. A new strategy employing MSG/ethanol mixed feeding improved biomass generation as well as recombinant protein yield. Cell density of 100–120 A600 units/ml was achieved after 72 h of induction in shake flask cultivations, resulting in recombinant protein yield from PPEPCK that is comparable or even higher than that from PAOX1.

Conclusions

We have designed an induction medium for recombinant protein production from K. phaffii PPEPCK in shake flask cultivations. It consists of 1.0% yeast extract, 2.0% peptone, 0.17% yeast nitrogen base with ammonium sulfate, 100 mM potassium phosphate (pH 6.0), 0.4 mg/L biotin, 2.0% MSG, and 2% ethanol. Substitution of ammonium sulphate with 0.5% urea is optional. Carbon source was replenished every 24 h during 72 h induction period. Under these conditions, GFP and RBD yields from PPEPCK equaled and even surpassed those from PAOX1. Compared to the traditional methanol-inducible expression system, the inducers of glutamate-inducible expression system are non-toxic and their metabolism does not generate toxic metabolites such as formaldehyde and hydrogen peroxide. This study sets the stage for MSG-inducible, industrial scale recombinant protein production from K. phaffii PPEPCK in bioreactors.
Genome-wide CRISPR screen identifies ESPL1 limits the response of gastric cancer cells to apatinib
Authors: Zhang Bei , Chen Yan , Chen Xinqi , Ren Zhiyao , Xiang Hong , Mao Lipeng , Zhu Guodong
Published: February 2024

Apatinib was the first anti-angiogenic agent approved for treatment of metastatic gastric cancer (GC). However, the emergence of resistance was inevitable. Thus investigating new and valuable off-target effect of apatinib directly against cancer cells is of great significance. Here, we identified extra spindle pole bodies-like 1 (ESPL1) was responsible for apatinib resistance in GC cells through CRISPR genome-wide gain-of-function screening. Loss of function studies further showed that ESPL1 inhibition suppressed cell proliferation, migration and promoted apoptosis in vitro, and accordingly ESPL1 knockdown sensitized GC cells to apatinib. In addition, we found ESPL1 interacted with mouse double minute 2 (MDM2), a E3 ubiquitin protein ligase, and the combination of MDM2 siRNA with apatinib synergistically ameliorated the resistance induced by ESPL1 overexpression. In summary, our study indicated that ESPL1 played a critical role in apatinib resistance in GC cells. Inhibition of MDM2 could rescue the sensitivity of GC cells to apatinib and reverse ESPL1-mediated resistance.
Emerging trends in the coexistence of primary lung Cancer and hematologic malignancy: a comprehensive analysis of clinicopathological features and genetic abnormalities
Authors: Lyu Mengchen , Luo Lifeng , Zhou Ling , Feng Xiangran , Yang Jin , Xu Ziwei , Sun Xianwen , Bao Zhiyao , Wang Xiaofei , Gao Beili , Xiang Yi
Published: February 2024

Background

The incidence of multiple primary cancers (MPC), especially involving primary lung cancer (PLC) and primary hematologic malignancies (PHM), is rising. This study aims to analyze clinicopathological features, gene abnormalities, and prognostic outcomes in individuals diagnosed with PLC-PHM MPC.

Methods

A retrospective analysis included 89 patients diagnosed with PLC-PHM MPC at the Respiratory or Hematology Departments of Ruijin Hospital from 2003 to 2022 (a total of 842,047 people). Next-generation sequencing (NGS) assessed lung cancer specimens, while Polymerase Chain Reaction (PCR) and NGS were used for hematologic malignancy specimens. Statistical analysis involved survival analysis and Cox regression.

Results

PLC-PHM MPC incidence surged from 1.67 per year (2011–2013) to 16.3 per year (2020–2022). The primary demographic for PLC-PHM MPC consists predominantly of elderly (average age 66 years) males (59.6%), with a high prevalence of metachronous MPC (89.9%). The prevailing histological types were lung adenocarcinoma (70.8%) in lung cancer (LC) and mature B-cell lymphomas (50.6%) in hematologic malignancies (HM). Notably, in a molecular testing cohort of 38 LC patients, 84.2% of lung cancer cases exhibited driver mutations, in which EGFR mutations frequence prevalent was 74.2%. In total group of 85 cases achieved a median overall survival (mOS) of 46.2 months, with a 5-year survival rate of 37.9% and advanced LC patients with LC gene mutations achieved a mOS was 52.6 months, with a 5-year OS rate of 30.6%. The median progression-free survival (PFS) following first-line treatment of 11 advanced patients with lung cancer-associated driver gene mutations is 26.6 months. Multivariate Cox regression revealed a favorable OS associated with surgery for LC, favorable PS score, adenocarcinoma pathology of LC, and the presence of genetic abnormalities associated with HM.

Conclusion

PLC-PHM MPC incidence is rising, characterized by a significant proportion of lung adenocarcinoma and a high prevalence of positive driver genes, especially in EGFR. Despite suffering from two primary tumors, the PLC-PHM MPC patients had superior data of both PFS and OS, suggesting an inherently intricate background of genetic abnormalities between the two kinds of tumors.
Molecular epidemiological characteristics of osteoarthritis-associated Brucella melitensis in China: evidence from whole-genome sequencing-based analysis
Authors: Zhu Lei , Zhang Chi , Liang Chen , Peng Li , Yan Huanyu , Liang Xiuwen , Xu Youjia
Published: February 2024

Background

Brucellosis, developing complications including arthritis, spondylitis, sacroiliitis, and osteomyelitis, is one of the most common zoonotic diseases in the current world which causes economic losses to the livestock industry and is a great public health concern. Brucella melitensis are the main pathogen of brucellosis epidemics in China, most of which are located in northern China. However, there is limited knowledge about the epidemiology of osteoarthritis-associated brucellosis. This study was aimed to reveal the prevalence of osteoarthritis-associated brucellosis in Inner Mongolia and also to investigate the molecular characteristics of B. melitensis isolates.

Methods and results

In 2018, the osteoarthritis symptoms of brucellosis in the Brucellosis department of a hospital in Inner Mongolia were investigated. Twenty osteoarthritis-associated B. melitensis strains, isolated from the inpatients in Inner Mongolia during 2013–2017, were subjected to whole genome sequencing. The multilocus sequence type (MLST) and core genome SNP (cgSNP) analysis were conducted to detect molecular epidemiological characteristics. The incidence of brucellosis osteoarthritis symptoms in males (85/120, 70.8%) was significantly higher than that in females (35/120, 29.2%), and the age of patients was concentrated between 41 and 60 years old. In silico analyses indicated ST8 was the prevalent sequence type and the transmission of osteoarthritis-associated B. melitensis among different geographical areas. All strains carry virulence genes, including cgs, lpsA, manCoAg, pgm, pmm, virB4, wbdA and wboA.

Conclusion

Our study showed the close epidemiologically connection of osteoarthritis-associated B. melitensis strains in northern China. And ST8 was the prevalent sequence type which need our attention.
Characterization of resistance genes and replicon typing in Carbapenem-resistant Klebsiella pneumoniae strains
Authors: Karaman Esin , Çiçek Ayşegül Çopur , Şemen Vicdan , Şaban Beriş Fatih
Published: February 2024

Objective

In our study, K. pneumoniae strains (non-susceptible to carbapenem) (n = 60) were obtained from various clinical samples from Rize State Hospital between 2015 and 2017 and it is aimed to identify antibiotic resistance genes and replicon typing.

Methods

Antibiotic susceptibility tests of the strains were performed with Kirby-Bauer disk diffusion test and the Vitek-2 automated system (BioMerieux, France). Antibiotic resistance genes and replicon typing was characterized by PCR method.

Results

It was determined that K. pneumaniae isolates were mostly isolated from the samples of the intensive care unit. All of the K. pneumoniae strains examined in this study were found to be ampicillin/sulbactam and ertapenem resistant but colistin susceptible. Amoxacillin/clavulonic acid resistance was detected at 98.14% of strains. The blaOXA-48 gene was mostly detected in isolates. The most common type of plasmid was I1 and 3 different plasmid types were found in five different strains together.

Conclusion

This study also shows that the distribution of NDM-1 and OXA-48 carbapenemases has increased since the first co-display in Türkiye and that IncHI1 is the first record in our country. This study provides an overview of the major plasmid families occurring in multiple antibiotic-resistant strains of K. pneumoniae. To our knowledge, this study represents the first report of IncHI1 record in Türkiye.
Correlation between antimicrobial resistance, biofilm formation, and virulence determinants in uropathogenic Escherichia coli from Egyptian hospital
Authors: Alshaikh Sara A. , El-banna Tarek , Sonbol Fatma , Farghali Mahmoud H.
Published: February 2024

Background

Uropathogenic Escherichia coli (UPEC) is the main etiological agent behind community-acquired and hospital-acquired urinary tract infections (UTIs), which are among the most prevalent human infections. The management of UPEC infections is becoming increasingly difficult owing to multi-drug resistance, biofilm formation, and the possession of an extensive virulence arsenal. This study aims to characterize UPEC isolates in Tanta, Egypt, with regard to their antimicrobial resistance, phylogenetic profile, biofilm formation, and virulence, as well as the potential associations among these factors.

Methods

One hundred UPEC isolates were obtained from UTI patients in Tanta, Egypt. Antimicrobial susceptibility was assessed using the Kirby-Bauer method. Extended-spectrum β-lactamases (ESBLs) production was screened using the double disk synergy test and confirmed with PCR. Biofilm formation was evaluated using the microtiter-plate assay and microscopy-based techniques. The phylogenetic groups of the isolates were determined. The hemolytic activity, motility, siderophore production, and serum resistance of the isolates were also evaluated. The clonal relatedness of the isolates was assessed using ERIC-PCR.

Results

Isolates displayed elevated resistance to cephalosporins (90–43%), sulfamethoxazole-trimethoprim (63%), and ciprofloxacin (53%). Ninety percent of the isolates were multidrug-resistant (MDR)/ extensively drug-resistant (XDR) and 67% produced ESBLs. Notably, there was an inverse correlation between biofilm formation and antimicrobial resistance, and 31%, 29%, 32%, and 8% of the isolates were strong, moderate, weak, and non-biofilm producers, respectively. Beta-hemolysis, motility, siderophore production, and serum resistance were detected in 64%, 84%, 65%, and 11% of the isolates, respectively. Siderophore production was correlated to resistance to multiple antibiotics, while hemolysis was more prevalent in susceptible isolates and associated with stronger biofilms. Phylogroups B2 and D predominated, with lower resistance and stronger biofilms in group B2. ERIC-PCR revealed considerable diversity among the isolates.

Conclusion

This research highlights the dissemination of resistance in UPEC in Tanta, Egypt. The evident correlation between biofilm and resistance suggests a resistance cost on bacterial cells; and that isolates with lower resistance may rely on biofilms to enhance their survival. This emphasizes the importance of considering biofilm formation ability during the treatment of UPEC infections to avoid therapeutic failure and/or infection recurrence.
Small cells – big issues: biological implications and preclinical advancements in small cell lung cancer
Authors: Solta Anna , Ernhofer Büsra , Boettiger Kristiina , Megyesfalvi Zsolt , Heeke Simon , Hoda Mir Alireza , Lang Christian , Aigner Clemens , Hirsch Fred R. , Schelch Karin , Döme Balazs
Published: February 2024

Current treatment guidelines refer to small cell lung cancer (SCLC), one of the deadliest human malignancies, as a homogeneous disease. Accordingly, SCLC therapy comprises chemoradiation with or without immunotherapy. Meanwhile, recent studies have made significant advances in subclassifying SCLC based on the elevated expression of the transcription factors ASCL1, NEUROD1, and POU2F3, as well as on certain inflammatory characteristics. The role of the transcription regulator YAP1 in defining a unique SCLC subset remains to be established. Although preclinical analyses have described numerous subtype-specific characteristics and vulnerabilities, the so far non-existing clinical subtype distinction may be a contributor to negative clinical trial outcomes. This comprehensive review aims to provide a framework for the development of novel personalized therapeutic approaches by compiling the most recent discoveries achieved by preclinical SCLC research. We highlight the challenges faced due to limited access to patient material as well as the advances accomplished by implementing state-of-the-art models and methodologies.
Activation-induced cytidine deaminase causes recurrent splicing mutations in diffuse large B-cell lymphoma
Authors: Benitez-Cantos Maria S. , Cano Carlos , Cuadros Marta , Medina Pedro P.
Published: February 2024

Diffuse large B-cell lymphoma (DLBCL) is the most common lymphoma. A major mutagenic process in DLBCL is aberrant somatic hypermutation (aSHM) by activation-induced cytidine deaminase (AID), which occurs preferentially at RCH/TW sequence motifs proximal to transcription start sites. Splice sequences are highly conserved, rich in RCH/TW motifs, and recurrently mutated in DLBCL. Therefore, we hypothesized that aSHM may cause recurrent splicing mutations in DLBCL. In a meta-cohort of > 1,800 DLBCLs, we found that 77.5% of splicing mutations in 29 recurrently mutated genes followed aSHM patterns. In addition, in whole-genome sequencing (WGS) data from 153 DLBCLs, proximal mutations in splice sequences, especially in donors, were significantly enriched in RCH/TW motifs (p < 0.01). We validated this enrichment in two additional DLBCL cohorts (N > 2,000; p < 0.0001) and confirmed its absence in 12 cancer types without aSHM (N > 6,300). Comparing sequencing data from mouse models with and without AID activity showed that the splice donor sequences were the top genomic feature enriched in AID-induced mutations (p < 0.0001). Finally, we observed that most AID-related splice site mutations are clonal within a sample, indicating that aSHM may cause early loss-of-function events in lymphomagenesis. Overall, these findings support that AID causes an overrepresentation of clonal splicing mutations in DLBCL.

Graphical Abstract
Crosstalk with renal proximal tubule cells drives acidosis-induced inflammatory response and dedifferentiation of fibroblasts via p38-singaling
Authors: Schulz Marie-Christin , Kopf Michael , Gekle Michael
Published: February 2024

Background

Tubulointerstitial kidney disease associated microenvironmental dysregulation, like acidification, inflammation and fibrosis, affects tubule cells and fibroblasts. Micromilieu homeostasis influences intracellular signaling and intercellular crosstalk. Cell–cell communication in turn modulates the interstitial microenvironment. We assessed the impact of acidosis on inflammatory and fibrotic responses in proximal tubule cells and fibroblasts as a function of cellular crosstalk. Furthermore, cellular signaling pathways involved were identified.

Methods

HK-2 (human proximal tubule) and CCD-1092Sk (human fibroblasts), in mono and coculture, were exposed to acidic or control media for 3 or 48 h. Protein expression of inflammation markers (TNF, TGF-ß and COX-2), dedifferentiation markers (N-cadherin, vinculin, ß-catenin and vimentin), fibrosis markers (collagen III and fibronectin) and phospho- as well as total MAPK levels were determined by western blot. Secreted collagen III and fibronectin were measured by ELISA. The impact of MAPK activation was assessed by pharmacological intervention. In addition, necrosis, apoptosis and epithelial permeability were determined.

Results

Independent of culture conditions, acidosis caused a decrease of COX-2, vimentin and fibronectin expression in proximal tubule cells. Only in monoculture, ß-Catenin expression decreased and collagen III expression increased in tubule cells during acidosis. By contrast, in coculture collagen III protein expression of tubule cells was reduced. In fibroblasts acidosis led to an increase of TNF, COX-2, vimentin, vinculin, N-cadherin protein expression and a decrease of TGF-ß expression exclusively in coculture. In monoculture, expression of COX-2 and fibronectin was reduced. Collagen III expression of fibroblasts was reduced by acidosis independent of culture conditions.

In coculture, acidosis enhanced phosphorylation of ERK1/2, JNK1/2 and p38 transiently in proximal tubule cells. In fibroblasts, acidosis enhanced phosphorylation of p38 in a sustained and very strong manner. ERK1/2 and JNK1/2 were not affected in fibroblasts. Inhibition of JNK1/2 and p38 under coculture conditions reduced acidosis-induced changes in fibroblasts significantly.

Conclusions

Our data show that the crosstalk between proximal tubule cells and fibroblasts is crucial for acidosis-induced dedifferentiation of fibroblasts into an inflammatory phenotype. This dedifferentiation is at least in part mediated by p38 and JNK1/2. Thus, cell–cell communication is essential for the pathophysiological impact of tubulointerstitial acidosis.
Apoptosis dysfunction: unravelling the interplay between ZBP1 activation and viral invasion in innate immune responses
Authors: Zhan Jianhao , Wang Jisheng , Liang Yuqing , Wang Lisha , Huang Le , Liu Shanshan , Zeng Xiaoping , Zeng Erming , Wang Hongmei
Published: February 2024

Apoptosis plays a pivotal role in pathogen elimination and maintaining homeostasis. However, viruses have evolved strategies to evade apoptosis, enabling their persistence within the host. Z-DNA binding protein 1 (ZBP1) is a potent innate immune sensor that detects cytoplasmic nucleic acids and activates the innate immune response to clear pathogens. When apoptosis is inhibited by viral invasion, ZBP1 can be activated to compensate for the effect of apoptosis by triggering an innate immune response. This review examined the mechanisms of apoptosis inhibition and ZBP1 activation during viral invasion. The authors outlined the mechanisms of ZBP1-induced type I interferon, pyroptosis and necroptosis, as well as the crosstalk between ZBP1 and the cGAS-STING signalling pathway. Furthermore, ZBP1 can reverse the suppression of apoptotic signals induced by viruses. Intriguingly, a positive feedback loop exists in the ZBP1 signalling pathway, which intensifies the innate immune response while triggering a cytokine storm, leading to tissue and organ damage. The prudent use of ZBP1, which is a double-edged sword, has significant clinical implications for treating infections and inflammation.
Inhibition of ferroptosis reverses heart failure with preserved ejection fraction in mice
Authors: Xiong Yixiao , Liu Xin , Jiang Ling , Hao Tao , Wang Yanyan , Li Tao
Published: February 2024

Background

Heart failure with preserved ejection fraction (HFpEF) accounts for approximately 50% of heart failure cases. The molecular mechanisms by which HFpEF leads to impaired diastolic function of the heart have not been clarified, nor have the drugs that target the clinical symptoms of HFpEF patients.

Methods

HFpEF chip data (GSE180065) was downloaded from the National Center for Biotechnology Information (NCBI) database. Differentially expressed genes (DEGs) were filtered by the limma package in R and processed for GO and KEGG pathway analyses. Then, ferroptosis-related genes in HFpEF were identified by taking the intersection between DEGs and ferroptosis-related genes. CytoHubba and MCODE were used to screen ferroptosis-related hub DEGs in the protein–protein interaction (PPI) network. Establishment of a mouse HFpEF model to validate the transcript levels of ferroptosis-related hub DEGs and ferroptosis-related phenotypes. Transcript levels of ferroptosis-related hub DEGs and HFpEF phenotypic changes in the hearts of HFpEF mice were further examined after the use of ferroptosis inhibitors.

Results

GO and KEGG enrichment analyses suggested that the DEGs in HFpEF were significantly enriched in ferroptosis-related pathways. A total of 24 ferroptosis-related DEGs were identified between the ferroptosis gene dataset and the DEGs. The established PPI network was further analyzed by CytoHubba and MCODE modules, and 11 ferroptosis-related hub DEGs in HFpEF were obtained. In animal experiments, HFpEF mice showed significant abnormal activation of ferroptosis. The expression trends of the 11 hub DEGs associated with ferroptosis, except for Cdh1, were consistent with the results of the bioinformatics analysis. Inhibition of ferroptosis alters the transcript levels of 11 ferroptosis-related hub DEGs and ameliorates HFpEF phenotypes.

Conclusions

The present study contributes to a deeper understanding of the specific mechanisms by which ferroptosis is involved in the development of HFpEF and suggests that inhibition of ferroptosis may mitigate the progression of HFpEF. In addition, eleven hub genes were recognized as potential drug binding targets.
Acute venous thromboembolism plasma and red blood cell metabolomic profiling reveals potential new early diagnostic biomarkers: observational clinical study
Authors: Febra Cláudia , Saraiva Joana , Vaz Fátima , Macedo João , Al-Hroub Hamza Mohammad , Semreen Mohammad Harb , Maio Rui , Gil Vitor , Soares Nelson , Penque Deborah
Published: February 2024

Background

Venous thromboembolism (VTE) is a leading cause of cardiovascular mortality. The diagnosis of acute VTE is based on complex imaging exams due to the lack of biomarkers. Recent multi-omics based research has contributed to the development of novel biomarkers in cardiovascular diseases. Our aim was to determine whether patients with acute VTE have differences in the metabolomic profile compared to non-acute VTE.

Methods

This observational trial included 62 patients with clinical suspicion of acute deep vein thrombosis or pulmonary embolism, admitted to the emergency room. There were 50 patients diagnosed with acute VTE and 12 with non-acute VTE conditions and no significant differences were found between the two groups for clinical and demographic characteristics. Metabolomics assays identified and quantified a final number of 91 metabolites in plasma and 55 metabolites in red blood cells (RBCs). Plasma from acute VTE patients expressed tendency to a specific metabolomic signature, with univariate analyses revealing 23 significantly different molecules between acute VTE patients and controls (p < 0.05). The most relevant metabolic pathway with the strongest impact on the acute VTE phenotype wasd-glutamine andd-glutamate (p = 0.001, false discovery rate = 0.06). RBCs revealed a specific metabolomic signature in patients with a confirmed diagnosis of DVT or PE that distinguished them from other acutely diseased patients, represented by 20 significantly higher metabolites and four lower metabolites. Three of those metabolites revealed high performant ROC curves, including adenosine 3′,5′-diphosphate (AUC 0.983), glutathione (AUC 0.923), and adenine (AUC 0.91). Overall, the metabolic pathway most impacting to the differences observed in the RBCs was the purine metabolism (p = 0.000354, false discovery rate = 0.68).

Conclusions

Our findings show that metabolite differences exist between acute VTE and nonacute VTE patients admitted to the ER in the early phases. Three potential biomarkers obtained from RBCs showed high performance for acute VTE diagnosis. Further studies should investigate accessible laboratory methods for the future daily practice usefulness of these metabolites for the early diagnosis of acute VTE in the ER.
Cytosolic Cadherin 4 promotes angiogenesis and metastasis in papillary thyroid cancer by suppressing the ubiquitination/degradation of β-catenin
Authors: Wu Luyao , Xiao Jian , Yi Dandan , Ding Haoran , Wang Ru , Duan Zehua , Liu Zhijian , Shi Xianbiao , Shen Meiping , Sang Jianfeng
Published: February 2024

Background

Although the long-term prognosis of papillary thyroid cancer (PTC) is favorable, distant metastasis significantly compromises the prognosis and quality of life for patients with PTC. The Cadherin family plays a pivotal role in tumor metastasis; however, the involvement of Cadherin 4 (CDH4) in the metastatic cascade remains elusive.

Methods

The expression and subcellular localization of CDH4 were determined through immunohistochemistry, immunofluorescence, and western blot analyses. The impact of CDH4 on cell migration, invasion, angiogenesis, and metastasis was assessed using transwell assays, tube formation assays, and animal experiments. Immunoprecipitation assay and mass spectrometry were employed to examine protein associations. The influence of CDH4 on the subcellular expression of β-catenin and active β-catenin was investigated via western blotting and immunofluorescence. Protein stability and ubiquitination assay were employed to verify the impact of CDH4 on β-catenin degradation. Rescue experiments were performed to ensure the significance of CDH4 in regulating nuclear β-catenin signaling.

Results

CDH4 was found to be significantly overexpressed in PTC tissues and predominantly localized in the cytoplasm. Furthermore, the overexpression of CDH4 in tumor tissues is associated with lymph node metastasis in PTC patients. Cytosolic CDH4 promoted the migration, invasion, and lung metastasis of PTC cells and stimulated the angiogenesis and tumorigenesis of PTC; however, this effect could be reversed by Tegavivint, an antagonist of β-catenin. Mechanistically, cytosolic CDH4 disrupted the interaction between β-catenin and β-TrCP1, consequently impeding the ubiquitination process of β-catenin and activating the nuclear β-catenin signaling.

Conclusions

CDH4 induces PTC angiogenesis and metastasis via the inhibition of β-TrCP1-dependent ubiquitination of β-Catenin.
Understanding mother-to-child transmission of HIV among mothers engaged in HIV care in Kenya: a case report
Authors: Tuthill Emily L. , Odhiambo Belinda C. , Maltby Ann E.
Published: February 2024

Background

Mother-to-child transmission of HIV, which may occur in utero, during birth, or through breastmilk, is now largely preventable with the advancement of HIV testing and treatment for women and their infants. Globally, great progress has been recorded over the years, with a 58% decline in new infections in children from 2010 to 2022. Currently, Kenya is among the countries with the highest rates of mother-to-child transmission of HIV despite consistent efforts to promote prevention of mother to child transmission strategies.

Methods

This case report presents the experiences of a woman, engaged in HIV care in Kenya, whose baby contracted HIV. The data used to describe this case come from surveys, provider notes, health records, observational notes, notes from phone call consultations, and one in-depth interview. All data sources were carefully reviewed, compared and complied to describe the timeline of events and context of the participant’s experience.

Results

We found multiple factors which may have contributed to this case of mother-to-child transmission of HIV. Antenatal care was initiated late in pregnancy (during the third trimester), and as a result, HIV diagnosis and treatment also occurred late in pregnancy. In addition, a lack of coordination between the clinic providing antenatal care and HIV treatment, and the hospital providing labor and delivery services led to breastfeeding initiation prior to the administration of infant HIV prophylaxis medications. Finally, poor maternal adherence to HIV medications went undetected and unaddressed until it was revealed by routine viral load monitoring three months after initiating HIV treatment (more than two months postpartum).

Conclusions

Our case report shows the continued need for more intensive and integrated care for mothers living with HIV and their infants including support for pregnant women newly diagnosed with HIV, coordination of perinatal and HIV care, provisions for routine monitoring of HIV medication adherence, intensive follow-up care including point of care testing for HIV exposed infants and in person breastfeeding support. Our case report contributes an important perspective especially in light of the current UNAIDS Global AIDS Strategy which recently inspired the Global Alliance to end AIDS in Children.
Mapping awareness-raising and capacity-building materials on developmental disabilities for non-specialists: a review of the academic and grey literature
Authors: Genovesi Elisa , Yao Yuan Ishtar , Mitchell Emily , Arad Michal , Diamant Victoria , Panju Areej , Hanlon Charlotte , Tekola Bethlehem , Hoekstra Rosa A.
Published: February 2024

Most children with developmental disabilities (DD), such as intellectual disabilities and autism, live in low- and middle-income countries (LMICs), where services are usually limited. Various governmental, non-governmental and research organisations in LMICs have developed awareness-raising campaigns and training and education resources on DD in childhood relevant to LMICs. This study aimed to comprehensively search and review freely available materials in the academic and grey literature, aimed at awareness raising, training and education on DD among non-specialist professionals and community members in LMICs. We consulted 183 experts, conducted key-word searches in five academic databases, four grey-literature databases and seventeen customised Google search engines. Following initial screening, we manually searched relevant systematic reviews and lists of resources and conducted forwards and backwards citation checks of included articles. We identified 7327 articles and resources after deduplication. We then used a rigorous multi-step screening process to select 78 training resources on DD relevant to LMICs, of which 43 aimed at informing and/or raising awareness DD, 16 highlighted specific strategies for staff in health settings and 19 in education settings. Our mapping analysis revealed that a wealth of materials is available for both global and local use, including comics, children’s books, flyers, posters, fact sheets, blogs, videos, websites pages, social media channels, handbooks and self-education guides, and training programmes or sessions. Twelve resources were developed for cross-continental or global use in LMICs, 19 were developed for and/or used in Africa, 23 in Asia, 24 in Latin America. Most resources were developed within the context where they were intended to be used. Identified gaps included a limited range of resources on intellectual disabilities, manuals for actively delivering training to staff in education settings and resources targeted at eastern European LMICs: future intervention development and adaptation efforts should address such gaps, to ensure capacity building materials exist for a sufficient variety of DD, settings and geographical areas. Beyond identifying these gaps, the value of the review lies in the compilation of summary tables of information on all freely available resources found, to support their selection and use in wider contexts. Information on the resource content, country of original development and copyright is provided to facilitate resource sharing and uptake.
FFAR2 expressing myeloid-derived suppressor cells drive cancer immunoevasion
Authors: Zhao Zeda , Qin Juliang , Qian Ying , Huang Chenshen , Liu Xiaohong , Wang Ning , Li Liqin , Chao Yuqing , Tan Binghe , Zhang Na , Qian Min , Li Dali , Liu Mingyao , Du Bing
Published: February 2024

Background

Emerging evidences suggest that aberrant metabolites contributes to the immunosuppressive microenvironment that leads to cancer immune evasion. Among tumor immunosuppressive cells, myeloid-derived suppressor cells (MDSCs) are pathologically activated and extremely immunosuppressive, which are closely associated with poor clinical outcomes of cancer patients. However, the correlation between MDSCs mediated immunosuppression and particular cancer metabolism remained elusive.

Methods

Spontaneous lung adenocarcinoma and subcutaneous mouse tumor models, gas chromatography–mass spectrometry (GC–MS) and immunofluorescence assay of patient-derived lung adenocarcinoma tissues, and flow cytometry, RNA sequencing and Western blotting of immune cells, were utilized.

Results

Metabolite profiling revealed a significant accumulation of acetic acids in tumor tissues from both patients and mouse model, which contribute to immune suppression and cancer progression significantly through free fatty acid receptor 2 (FFAR2). Furthermore, FFAR2 is highly expressed in the myeloid-derived suppressor cells (MDSCs) from the tumor of lung adenocarcinoma (LUAD) patients which is greatly associated with poor prognosis. Surprisingly, whole or myeloid Ffar2 gene deletion markedly inhibited urethane-induced lung carcinogenesis and syngeneic tumor growth with reduced MDSCs and increased CD8+ T cell infiltration. Mechanistically, FFAR2 deficiency in MDSCs significantly reduced the expression of Arg1 through Gαq/Calcium/PPAR-γ axis, which eliminated T cell dysfunction through relieving L-Arginine consumption in tumor microenvironment. Therefore, replenishment of L-Arginine or inhibition to PPAR-γ restored acetic acids/FFAR2 mediated suppression to T cells significantly. Finally, FFAR2 inhibition overcame resistance to immune checkpoint blockade through enhancing the recruitment and cytotoxicity of tumor-infiltrating T cells.

Conclusion

Altogether, our results demonstrate that the acetic acids/FFAR2 axis enhances MDSCs mediated immunosuppression through Gαq/calcium/PPAR-γ/Arg1 signaling pathway, thus contributing to cancer progression. Therefore, FFAR2 may serve as a potential new target to eliminate pathologically activated MDSCs and reverse immunosuppressive tumor microenvironment, which has great potential in improving clinical outcomes of cancer immunotherapy.
Effect of repeated vitrification of human embryos on pregnancy and neonatal outcomes
Authors: Huang Yan , Cheng Yi , Zhang Min , Chen Yiqing , Zhou Rong , Lin Dewei , Guo Xinyu
Published: February 2024

Background

Repeated cryopreservation of embryos should occasionally be considered when embryos were not suitable for transfer. The effect of re-cryopreservation on embryos remains contentious.

Methods

This retrospective cohort study aimed to evaluate the pregnancy and neonatal outcomes of twice vitrificated blastocyst derived from once vitrified embryos. Total 410 vitrified-warmed blastocyst transfer cycles were divided into two groups according to the times of embryo vitrification: (1) vitrified blastocysts derived from fresh blastocysts (control group, n = 337); (2) twice vitrified blastocysts derived from once vitrified embryos (n = 73). The primary outcome was live birth rate. Multivariable logistic or linear regression analysis model was performed to describe the association between embryo cryopreservation times and clinical outcomes.

Results

No difference was observed in female age at retrieval and transfer, infertility period, body mass index (BMI), infertility type, endometrial thickness, and embryo transfer numbers between the two groups. The pregnancy outcomes of embryos in repeated cryopreservation group were comparable to those of embryos in control group, including implantation rate, clinical pregnancy rate, and live birth rate. In multivariate logistic regression analysis, the cryopreservation times did not affect the outcomes of biochemical pregnancy, clinical pregnancy, and live birth. Moreover, there was no difference in gestational age, birthweight and sex ratio of singleton newborns between groups. After correcting several possible confounding variables, no significant association was observed between cryopreservation times and neonatal birthweight.

Conclusion

In conclusion, pregnancy and neonatal outcomes achieved with twice vitrified blastocyst transfer were comparable to those achieved with vitrified blastocyst transfer in control group.
Comparing the effects of pulsed and radiofrequency catheter ablation on quality of life, anxiety, and depression of patients with paroxysmal supraventricular tachycardia: a single-center, randomized, single-blind, standard-controlled trial
Authors: Du Ying , Ma Shanshan , Yue Pan , Xu Ying , Wen Ya , Ji Mingzhu , He Lingxiao , Liao Dengbin
Published: February 2024

Background

Radiofrequency catheter ablation (RFCA) may lead to decreased quality of life (QOL) and increased anxiety and depression in patients with paroxysmal supraventricular tachycardia (PSVT), possibly due to the lack of selectivity of the ablation tissue and the long ablation time. In recent years, pulsed field ablation (PFA) has been used for the first time in China to treat PSVT patients because of its ability to ablate abnormal tissue sites in a precise and transient manner. This study was conducted to compare the effects of PFA and RFCA on QOL and psychological symptoms of PSVT patients.

Methods

We have designed a single-center, randomized, single-blind, standard-controlled trial. A total of 50 participants who met the eligibility criteria would be randomly allocated into the PFA group or RFCA group in a 1:1 ratio. All participants were assessed using the 36-Item Short-Form Health Survey (SF-36) and the Hospital Anxiety and Depression Scale (HADS) at pre-procedure (T0), post-procedure (T1), and 3 months post-procedure (T2). The SPSS 21.0 software was used to analyze the data through Wilcoxon and Fisher’s exact tests and repeated measures ANOVA.

Results

Twenty-five in the PFA group and 24 in the RFCA group completed the trial. SF-36: (1) Between-group comparison: At T1, PFA group had significantly higher SF-36 scores on physiological function (PF) and general health (GH) than RFCA group, with a treatment difference of 5.61 points and 18.51 points(P < 0.05). (2) Within-group comparison: We found that in the PFA and RFCA groups, T2 showed significant improvement in the remaining 6 subscales of the SF-36 scale compared to T1 and T0 (P < 0.05), except for body pain (BP) and social function (SF) scores. HADS: (1) Between-group comparison: no significant difference (P > 0.05). (2) Within-group comparison: The HADS scores of the PFA and RFCA groups were statistically significant at T2 compared to T0 and T1 (P < 0.05).

Conclusions

Our study provided new and meaningful evidence that PFA was effective in significantly improving QOL and decreasing anxiety and depression in PFA patients.

Trial registration

Chinese Clinical Trial Registry, ChiCTR2200060272.