bluuebunny/crossref_metadata_2025 huggingface dataset:
Recombinant production of Paenibacillus wynnii β-galactosidase with Komagataella phaffii
Authors: Bechtel Anna , Seitl Ines , Pross Eva , Hetzel Frank , Keutgen Mario , Fischer Lutz
Published: October 2024

Background

The β-galactosidase from Paenibacillus wynnii (β-gal-Pw) is a promising candidate for lactose hydrolysis in milk and dairy products, as it has a higher affinity for the substrate lactose (low KM value) compared to industrially used β-galactosidases and is not inhibited by the hydrolysis-generated product D-galactose. However, β-gal-Pw must firstly be produced cost-effectively for any potential industrial application. Accordingly, the yeast Komagataella phaffii was chosen to investigate its feasibility to recombinantly produce β-gal-Pw since it is approved for the regulated production of food enzymes. The aim of this study was to find the most suitable way to produce the β-gal-Pw in K. phaffii either extracellularly or intracellularly.

Results

Firstly, 11 different signal peptides were tested for extracellular production of β-gal-Pw by K. phaffii under the control of the constitutive GAP promoter. None of the signal peptides resulted in a secretion of β-gal-Pw, indicating problems within the secretory pathway of this enzyme. Therefore, intracellular β-gal-Pw production was investigated using the GAP or methanol-inducible AOX1 promoter. A four-fold higher volumetric β-galactosidase activity of 7537 ± 66 µkatoNPGal/Lculture was achieved by the K. phaffii clone 27 using the AOX1 promoter in fed-batch bioreactor cultivations, compared to the clone 5 using the GAP promoter. However, a two-fold higher specific productivity of 3.14 ± 0.05 µkatoNPGal/gDCW/h was achieved when using the GAP promoter for β-gal-Pw production compared to the AOX1 promoter. After partial purification, a β-gal-Pw enzyme preparation with a total β-galactosidase activity of 3082 ± 98 µkatoNPGal was obtained from 1 L of recombinant K. phaffii culture (using AOX1 promoter).

Conclusion

This study showed that the β-gal-Pw was produced intracellularly by K. phaffii, but the secretion was not achieved with the signal peptides chosen. Nevertheless, a straightforward approach to improve the intracellular β-gal-Pw production with K. phaffii by using either the GAP or AOX1 promoter in bioreactor cultivations was demonstrated, offering insights into alternative production methods for this enzyme.
High-resolution spatio-temporal risk mapping for malaria in Namibia: a comprehensive analysis
Authors: Zhang Song , Amratia Punam , Symons Tasmin L. , Rumisha Susan F. , Kang Su Yun , Connell Mark , Uusiku Petrina , Katokele Stark , Hamunyela Jerobeam , Ntusi Nelly , Soroses Wilma , Moyo Ernest , Lukubwe Ophilia , Maponga Chivimbiso , Lucero Dominic , Gething Peter W. , Cameron Ewan
Published: October 2024

Background

Namibia, a low malaria transmission country targeting elimination, has made substantial progress in reducing malaria burden through improved case management, widespread indoor residual spraying and distribution of insecticidal nets. The country's diverse landscape includes regions with varying population densities and geographical niches, with the north of the country prone to periodic outbreaks. As Namibia approaches elimination, malaria transmission has clustered into distinct foci, the identification of which is essential for deployment of targeted interventions to attain the southern Africa Elimination Eight Initiative targets by 2030. Geospatial modelling provides an effective mechanism to identify these foci, synthesizing aggregate routinely collected case counts with gridded environmental covariates to downscale case data into high-resolution risk maps.

Methods

This study introduces innovative infectious disease mapping techniques to generate high-resolution spatio-temporal risk maps for malaria in Namibia. A two-stage approach is employed to create maps using statistical Bayesian modelling to combine environmental covariates, population data, and clinical malaria case counts gathered from the routine surveillance system between 2018 and 2021.

Results

A fine-scale spatial endemicity surface was produced for annual average incidence, followed by a spatio-temporal modelling of seasonal fluctuations in weekly incidence and aggregated further to district level. A seasonal profile was inferred across most districts of the country, where cases rose from late December/early January to a peak around early April and then declined rapidly to a low level from July to December. There was a high degree of spatial heterogeneity in incidence, with much higher rates observed in the northern part and some local epidemic occurrence in specific districts sporadically.

Conclusions

While the study acknowledges certain limitations, such as population mobility and incomplete clinical case reporting, it underscores the importance of continuously refining geostatistical techniques to provide timely and accurate support for malaria elimination efforts. The high-resolution spatial risk maps presented in this study have been instrumental in guiding the Namibian Ministry of Health and Social Services in prioritizing and targeting malaria prevention efforts. This two-stage spatio-temporal approach offers a valuable tool for identifying hotspots and monitoring malaria risk patterns, ultimately contributing to the achievement of national and sub-national elimination goals.
Perceived discrimination and refraining from seeking physician’s care in Sweden: an intersectional analysis of individual heterogeneity and discriminatory accuracy (AIHDA)
Authors: Hassan Mariam , Öberg Johan , Wemrell Maria , Vicente Raquel Perez , Lindström Martin , Merlo Juan
Published: October 2024

Background

Discrimination may further impede access to medical care for individuals in socially disadvantaged positions. Sociodemographic information and perceived discrimination intersect and define multiple contexts or strata that condition the risk of refraining from seeking physician’s care. By applying analysis of individual heterogeneity and discriminatory accuracy (AIHDA) we aimed to improve the mapping of risk by considering both strata average risk differences and the accuracy of such strata risks for distinguishing between individuals who did or did not refrain from seeking physician’s care.

Methods

We analysed nine annual National Public Health Surveys (2004, 2007–2014) in Sweden including 73,815 participants. We investigated the risk of refraining from seeking physician’s care across 64 intersectional strata defined by sex, education, age, country of birth, and perceived discrimination. We calculated strata-specific prevalences and prevalence ratios (PR) with 95% confidence intervals (CI), and the area under the receiver operating characteristic curve (AUC) to evaluate the discriminatory accuracy (DA).

Results

Discriminated foreign-born women aged 35–49 with a low educational level show a six times higher risk (PR = 6.07, 95% CI 5.05–7.30) than non-discriminated native men with a high educational level aged 35–49. However, the DA of the intersectional strata was small (AUC = 0.64). Overall, discrimination increased the absolute risk of refraining from seeking physician’s care, over and above age, sex, and educational level.

Conclusions

AIHDA disclosed complex intersectional inequalities in the average risk of refraining from seeking physician’s care. This risk was rather high in some strata, which is relevant from an individual perspective. However, from a population perspective, the low DA of the intersectional strata suggests that potential interventions to reduce such inequalities should be universal but tailored to the specific contextual characteristics of the strata. Discrimination impairs access to healthcare.
Pharmacological targeting of P300/CBP reveals EWS::FLI1-mediated senescence evasion in Ewing sarcoma
Authors: Wei Erdong , Mitanoska Ana , O’Brien Quinn , Porter Kendall , Molina MacKenzie , Ahsan Haseeb , Jung Usuk , Mills Lauren , Kyba Michael , Bosnakovski Darko
Published: October 2024

Ewing sarcoma (ES) poses a significant therapeutic challenge due to the difficulty in targeting its main oncodriver, EWS::FLI1. We show that pharmacological targeting of the EWS::FLI1 transcriptional complex via inhibition of P300/CBP drives a global transcriptional outcome similar to direct knockdown of EWS::FLI1, and furthermore yields prognostic risk factors for ES patient outcome. We find that EWS::FLI1 upregulates LMNB1 via repetitive GGAA motif recognition and acetylation codes in ES cells and EWS::FLI1-permissive mesenchymal stem cells, which when reversed by P300 inhibition leads to senescence of ES cells. P300-inhibited senescent ES cells can then be eliminated by senolytics targeting the PI3K signaling pathway. The vulnerability of ES cells to this combination therapy suggests an appealing synergistic strategy for future therapeutic exploration.
Proteomic analysis and experimental validation reveal the blood–brain barrier protective of Huanshaodan in the treatment of SAMP8 mouse model of Alzheimer’s disease
Authors: Su Yunfang , Liu Ningning , Wang Pan , Shang Congcong , Sun Ruiqin , Ma Jinlian , Li Zhonghua , Ma Huifen , Sun Yiran , Zhang Zijuan , Song Junying , Xie Zhishen , Xu Jiangyan , Zhang Zhenqiang
Published: October 2024

Background

Huanshaodan (HSD) is a Chinese Herbal Compound which has a definite clinical effect on Alzheimer’s disease (AD), however, the underlying mechanism remains unclear. The aim of this study is to preliminarily reveal the mechanism of HSD in the treatment of AD model of SAMP8 mice.

Methods

Chemical composition of HSD and its drug-containing serum were identified by Q-Orbitrap high resolution liquid mass spectrometry. Six-month-old SAMP8 mice were treated with HSD and Donepezil hydrochloride by gavage for 2 months, and Wogonin for 28 days. Behavioral test was performed to test the learning and memory ability of mice. Immunofluorescence (IF) or Western-blot methods were used to detect the levels of pSer404-tau and β-amyloid (Aβ) in the brain of mice. Hematoxylin–eosin (H&E) staining and Transmission electron microscopy (TEM) assay was applied to observe the pathological changes of neurons. Proteomic technology was carried out to analyze and identify the protein network of HSD interventions in AD. Then the pathological process of the revealed AD-related differential proteins was investigated by IF, Q-PCR, Western-blot, Fluorescence in situ hybridization (FISH) and 16S rRNA sequencing methods.

Results

The results showed that HSD and Wogonin, one of the components in its drug-containing serum, can effectively improve the cognitive impairments of SAMP8 mice, protect hippocampal neurons and synapses, and reduce the expression of pSer404-tau and Aβ. HSD and Wogonin reduced the levels of fibrinogen β chain (FGB) and γ chain (FGG), the potential therapeutic targets revealed by proteomics analysis, reduced the colocalization of FGB and FGG with Aβ, ionized calcium binding adaptor molecule 1 (Iba-1), glial fibrillary acidic protein (GFAP), increased level of and myelin basic protein (MBP). Meanwhile, HSD and Wogonin increased ZO-1 and Occludin levels, improved brain microvascular injury, and reduced levels of bacteria/bacterial DNA and lipopolysaccharide (LPS) in the brain of mice. In addition, 16S rRNA sequencing indicated that HSD regulated the structure of intestinal microbiota of mice.

Conclusion

The effects of HSD on AD may be achieved by inhibiting the levels of fibrinogen and the interactions on glia cells in the brain, and by modulating the structure of intestinal microbiota and improving the blood–brain barrier function.
Development of a new paradigm model for deciphering action mechanism of Danhong injection using a combination of isothermal shift assay and database interrogation
Authors: Wang Tianxiang , Yang Changmei , Tang Yuxiang , Wen Ke , Ma Yuxin , Chen Yuling , Li Zhiqiang , Zhao Yujiao , Zhu Songbiao , Meng Xianbin , Du Sijing , Miao Zelong , Wei Wei , Deng Haiteng
Published: October 2024

Background

Identification of active components of traditional Chinese Medicine (TCM) and their respective targets is important for understanding the mechanisms underlying TCM efficacy. However, there are still no effective technical methods to achieve this.

Methods

Herein, we have established a method for rapidly identifying targets of a specific TCM and interrogating the targets with their corresponding active components based on Isothermal Shift Assay (iTSA) and database interrogation.

Results

We optimized iTSA workflow and identified 110 targets for Danhong injection (DHI) which is used as an effective remedy for cardiovascular and cerebrovascular diseases. Moreover, we identified the targets of the nine major ingredients found in DHI. Database interrogation found that the potential targets for DHI, in which we verified that ADK as the target for salvianolic acid A and ALDH1B1 as the target for protocatechualdehyde in DHI, respectively.

Conclusion

Overall, we established a novel paradigm model for the identification of targets and their respective ingredients in DHI, which facilitates the discovery of drug candidates and targets for improving disease management and contributes to revealing the underlying mechanisms of TCM and fostering TCM development and modernization.

Graphical Abstract
Differences in seasonal dynamics and pyrethroid resistance development among Anopheles Hyrcanus group species
Authors: Lee Do Eun , Han Jeong Heum , Lee Gang Chan , Choi Junhyeong , Kwun Wonyong , Lee Si Hyeock , Kim Ju Hyeon
Published: October 2024

Background

The Anopheles Hyrcanus group, which transmits Plasmodium vivax, consists of six confirmed species in South Korea. An epidemiological study revealed differences in the seasonal occurrence patterns of each species. Pyrethroid resistance in An. sinensis dates back to the early 2000s, whereas information on pyrethroid resistance in other species is lacking despite their greater significance in malaria epidemiology.

Methods

Anopheles mosquitoes were collected from two malaria-endemic regions in South Korea for 2 years and their knockdown resistance (kdr) mutations were genotyped. The larval susceptibility to λ-cyhalothrin was compared in six Anopheles species and its seasonal changes in three species were investigated. The full-length sequences of the voltage-sensitive sodium channel (VSSC) were compared across six species to evaluate potential target-site insensitivity. The contribution of the kdr mutation to phenotypic resistance was confirmed by comparing median lethal time (LT50) to λ-cyhalothrin between populations of Anopheles belenrae with distinct genotypes.

Results

The composition and seasonal occurrence of rare species (Anopheles kleini, Anopheles lestri, and Anopheles sineroides) varied considerably, whereas An. sinensis occurs continuously throughout the season. A kdr mutation in the form of heterozygous allele was newly identified in An. belenrae, An. lesteri, An. pullus, and An. sineroides. The baseline susceptibility to λ-cyhalothrin was the highest in An. belenrae, followed by An. lesteri, An. sineroides, An. kleini, An. pullus, and An. sinensis, with median lethal concentration (LC50) values ranging from 6.0- to 73.5-fold higher than that of An. belenrae. The susceptibility of An. sinensis and An. pullus varied by season, whereas that of An. belenrae remained stable. The kdr-heterozygous An. belenare population exhibited 5.1 times higher LT50 than that of the susceptible population. Species-specific VSSC sequence differences were observed among the six species.

Conclusions

Our findings suggest that the status and extent of pyrethroid resistance vary among Anopheles Hyrcanus group species. While An. sinensis, the predominant species, developed a considerable level of pyrethroid resistance through kdr mutation, the resistance levels of other species appeared to be less pronounced. Large-scale monitoring is crucial to fully understand species-specific seasonal occurrence and resistance status for effective management strategies, considering the ongoing impact of climate change on their vectorial capacity.

Graphical Abstract
The goat pan-genome reveals patterns of gene loss during domestication
Authors: Liu Jiaxin , Shi Yilong , Mo Dongxin , Luo Lingyun , Xu Songsong , Lv Fenghua
Published: October 2024

Background

Unveiling genetic diversity features and understanding the genetic mechanisms of diverse goat phenotypes are pivotal in facilitating the preservation and utilization of these genetic resources. However, the total genetic diversity within a species can’t be captured by the reference genome of a single individual. The pan-genome is a collection of all the DNA sequences that occur in a species, and it is expected to capture the total genomic diversity of the specific species.

Results

We constructed a goat pan-genome using map-to-pan assemble based on 813 individuals, including 723 domestic goats and 90 samples from their wild relatives, which presented a broad regional and global representation. In total, 146 Mb sequences and 974 genes were identified as absent from the reference genome (ARS1.2; GCF_001704415.2). We identified 3,190 novel single nucleotide polymorphisms (SNPs) using the pan-genome analysis. These novel SNPs could properly reveal the population structure of domestic goats and their wild relatives. Presence/absence variation (PAV) analysis revealed gene loss and intense negative selection during domestication and improvement.

Conclusions

Our research highlights the importance of the goat pan-genome in capturing the missing genetic variations. It reveals the changes in genomic architecture during goat domestication and improvement, such as gene loss. This improves our understanding of the evolutionary and breeding history of goats.
Metaproteogenomics resolution of a high-CO2 aquifer community reveals a complex cellular adaptation of groundwater Gracilibacteria to a host-dependent lifestyle
Authors: Figueroa-Gonzalez Perla Abigail , Bornemann Till L. V. , Hinzke Tjorven , Maaß Sandra , Trautwein-Schult Anke , Starke Joern , Moore Carrie J. , Esser Sarah P. , Plewka Julia , Hesse Tobias , Schmidt Torsten C. , Schreiber Ulrich , Bor Batbileg , Becher Dörte , Probst Alexander J.
Published: October 2024

Background

Bacteria of the candidate phyla radiation (CPR), constituting about 25% of the bacterial biodiversity, are characterized by small cell size and patchy genomes without complete key metabolic pathways, suggesting a symbiotic lifestyle. Gracilibacteria (BD1-5), which are part of the CPR branch, possess alternate coded genomes and have not yet been cultivated. The lifestyle of Gracilibacteria, their temporal dynamics, and activity in natural ecosystems, particularly in groundwater, has remained largely unexplored. Here, we aimed to investigate Gracilibacteria activity in situ and to discern their lifestyle based on expressed genes, using the metaproteogenome of Gracilibacteria as a function of time in the cold-water geyser Wallender Born in the Volcanic Eifel region in Germany.

Results

We coupled genome-resolved metagenomics and metaproteomics to investigate a cold-water geyser microbial community enriched in Gracilibacteria across a 12-day time-series. Groundwater was collected and sequentially filtered to fraction CPR and other bacteria. Based on 725 Gbps of metagenomic data, 1129 different ribosomal protein S3 marker genes, and 751 high-quality genomes (123 population genomes after dereplication), we identified dominant bacteria belonging to Gallionellales and Gracilibacteria along with keystone microbes, which were low in genomic abundance but substantially contributing to proteomic abundance. Seven high-quality Gracilibacteria genomes showed typical limitations, such as limited amino acid or nucleotide synthesis, in their central metabolism but no co-occurrence with potential hosts. The genomes of these Gracilibacteria were encoded for a high number of proteins involved in cell to cell interaction, supporting the previously surmised host-dependent lifestyle, e.g., type IV and type II secretion system subunits, transporters, and features related to cell motility, which were also detected on protein level.

Conclusions

We here identified microbial keystone taxa in a high-CO2 aquifer, and revealed microbial dynamics of Gracilibacteria. Although Gracilibacteria in this ecosystem did not appear to target specific organisms in this ecosystem due to lack of co-occurrence despite enrichment on 0.2-µm filter fraction, we provide proteomic evidence for the complex machinery behind the host-dependent lifestyle of groundwater Gracilibacteria.
Ozone therapy mitigates parthanatos after ischemic stroke
Authors: Li Jiahui , Liu Xiaolei , Wang Zengze , Xie Pengyun , Zhu Min , Zhong Hanhui , Luo Sirui , Tang Jing , Mo Guixi
Published: October 2024

Background

Stroke is a leading cause of death worldwide, with oxidative stress and calcium overload playing significant roles in the pathophysiology of the disease. Ozone, renowned for its potent antioxidant properties, is commonly employed as an adjuvant therapy in clinical settings. Nevertheless, it remains unclear whether ozone therapy on parthanatos in cerebral ischemia–reperfusion injury (CIRI). This study aims to investigate the impact of ozone therapy on reducing parthanatos during CIRI and to elucidate the underlying mechanism.

Methods

Hydrogen peroxide (H2O2) was utilized to mimic the generation of reactive oxygen species (ROS) in SH-SY5Y cell reperfusion injury in vitro, and an in vivo ischemic stroke model was established. Ozone saline was introduced for co-culture or intravenously administered to mice. Apoptosis and oxidative stress were assessed using flow cytometry and immunofluorescence. Western blotting was utilized to examine the expression of parthanatos signature proteins. The mechanism by which ozone inhibits parthanatos was elucidated through inhibiting PPARg or Nrf2 activity.

Results

The findings demonstrated that ozone mitigated H2O2-induced parthanatos by either upregulating nuclear factor erythroid 2-related factor 2 (Nrf2) or activating peroxisome proliferator-activated receptorg (PPARg). Furthermore, through the use of calcium chelators and ROS inhibitors, it was discovered that ROS directly induced parthanatos and facilitated intracellular calcium elevation. Notably, a malignant feedback loop between ROS and calcium was identified, further amplifying the induction of parthanatos. Ozone therapy exhibited its efficacy by increasing PPARg activity or enhancing the Nrf2 translation, thereby inhibiting ROS production induced by H2O2. Concurrently, our study demonstrated that ozone treatment markedly inhibited parthanatos in stroke-afflicted mice. Additionally, ozone therapy demonstrated significant neuroprotective effects on cortical neurons, effectively suppressing parthanatos.

Conclusions

These findings contribute valuable insights into the potential of ozone therapy as a therapeutic strategy for reducing parthanatos during CIRI, highlighting its impact on key molecular pathways associated with oxidative stress and calcium regulation.
Planning for complex inferior vena cava filter retrievals: the implementation and effectiveness of 3D printed models
Authors: Lee Joonhyuk , Rybicki Frank J. , Ravi Prashanth , Chadalavada Seetharam C.
Published: October 2024

Background

Inferior vena cava filter (IVC) retrieval is most often routine but can be challenging with high morbidity in complex cases, especially those with an extended dwelling time. While risk of morbidity in complex retrievals is decreased with advanced filter retrieval techniques, deciding when and which to use these requires detailed pre-procedural planning. The purpose of our study was to evaluate patient-specific 3D printed anatomic IVC filter models for aiding complex IVC filter retrievals.

Methods

All IVC filter retrieval patients between June 2021 and September 2022 at one academic medical hospital were prospectively screened. Nine met criteria for complex retrieval, and their CT images were used to 3D print patient-specific IVC and filter models. Models were used in pre-procedural planning and clinical utility was assessed using the Anatomic Model Utility Likert Questionnaire and estimations of the procedural and fluoroscopy time saved.

Results

The usage of 3D printed models in pre-procedural planning had high clinical utility based on the Likert questionnaire (Anatomic Model Utility Points 366.7 ± 103.1). Using a model significantly increased confidence in planning (p = 0.03) and modified the treatment plan in seven cases. It also led to cost-efficient use of resources in the procedure suite with estimated reduction in procedure and fluoroscopy time of 29.0 [20.3] (p = 0.003) and 10.2 [6.7] (p = 0.002) minutes, respectively.

Conclusion

3D printed anatomic models for patients who require complex IVC filter retrieval demonstrated Likert-based high clinical utility and led to estimated reductions of procedural and fluoroscopy time.
Physical, functional and bioactive properties of microencapsulated powders from banana pseudostem and inflorescence extracts
Authors: Gayathry K. S. , John Jenny Ann
Published: October 2024

Application of agricultural by-products in the functional food and beverage industry is currently gaining prominence. Banana (Musa spp) is a popular tropical fruit with global production of 124.97 million tonnes. The banana production industry contributes to large amount of solid waste/ banana by-products, such as, pseudostem and inflorescence. Palayankodan (Musa × paradisiaca Mysore AAB group), Nendran (Musa × paradisiaca AAB group) and Njalipoovan (Musa × paradisiaca AB group) are three popular and common cultivars in Kerala, a state in South India. The present study was aimed to extract the potential bioactive compounds from the pseudostem and inflorescence of the above-mentioned cultivars and to standardise the process of microencapsulation using spray drying. Ultrasonication assisted extraction using ethanol as solvent was carried out. The extract and wall material parameters were standardised for microencapsulation. The encapsulated powders were analysed for encapsulation yield, retention efficiency, physical properties, phytochemical composition, antioxidant potential and anti-diabetic activity. The encapsulation yields and retention efficiencies of the encapsulated powders were found to be approximately 75 and 70.51%, respectively. Results revealed that the powders exhibited lower bulk density, good solubility and reconstitutability. Scanning electron microscopy was also conducted to reveal the particle morphology. All the powders exhibited smooth, spherical shape, with no pores. The phenolic and flavonoid contents of the encapsulated pseudostem and inflorescence extract powders ranged from 2.75 to 3.13 mg GAE/ g of powder and 34.83 to 46.67 mg QE/g of powder, respectively. The present study also reported the in vitro bioactive properties, in terms of antioxidant and anti-diabetic activities of the encapsulated powders. The HPLC analysis of the microencapsulated powders revealed the presence of gallic acid, protocatechuic acid, trans cinnamic acid, trans ferulic acid, epicatechin and syringic acid. It is clear from the study that the encapsulated powders from banana by-products have a great potential to be utilised by the functional food industry.

Graphical Abstract
7624 m6A mRNA Methylation in Brown Adipose Tissue Regulates Systemic Insulin Sensitivity via an Inter-Organ Prostaglandin Signaling Axis
Authors: Xiao Ling , De Jesus Dario F , Ju Cheng-Wei , Wei Jiang-bo , Hu Jiang , DiStefano-Forti Ava , Tsuji Tadataka , Cero Cheryl , Männistö Ville , Manninen Suvi M , Wei Si-Ying , Ijaduola Oluwaseun , Blüher Matthias , Cypess Aaron Martin , Pihlajamäki Jussi , Tseng Yu-Hua , He Chuan , Kulkarni Rohit N
Published: October 2024

Disclosure: L. Xiao: None. D. F De Jesus: None. C. Ju: None. J. Wei: None. J. Hu: None. A. DiStefano-Forti: None. T. Tsuji: None. C. Cero: None. V. Männistö: None. S.M. Manninen: None. S. Wei: None. O. Ijaduola: None. M. Blüher: Consulting Fee; Self; Amgen Inc, AstraZeneca, Bayer, Inc., Boehringer Ingelheim, Lilly USA, LLC, Novo Nordisk, Pfizer, Inc., Sanofi. A.M. Cypess: None. J. Pihlajamäki: None. Y. Tseng: None. C. He: Advisory Board Member; Self; Accent Therapeutics. R.N. Kulkarni: Advisory Board Member; Self; Novo Nordisk, Biomea, Inversago, REDD.

Brown adipose tissue (BAT) has the capacity to regulate systemic metabolism through the secretion of “Batokines” including signaling lipids. N6-methyladenosine (m6A) is the most prevalent and abundant post-transcriptional mRNA modification and has been reported to regulate BAT adipogenesis and energy expenditure. Here, we demonstrate that the absence of the m6A writer, methyltransferase-like 14 (METTL14), modifies the BAT secretome to initiate inter-organ communication to improve systemic insulin sensitivity. Importantly, these phenotypes are independent of UCP1-mediated energy expenditure and thermogenesis. Using lipidomics, we identified prostaglandin E2 (PGE2) and prostaglandin F2a (PGF2a) as M14KO-BAT-secreted insulin sensitizers. Notably, circulatory PGE2 and PGF2a levels are inversely correlated with insulin sensitivity in humans. Furthermore, in vivo administration of PGE2 and PGF2a in high-fat diet-induced insulin-resistant obese mice recapitulates the phenotypes of Mettl14-deficient animals. PGE2 or PGF2a improves insulin signaling by suppressing the expression of specific AKT phosphatases. Mechanistically, METTL14-mediated m6A installation promotes decay of transcripts encoding prostaglandin synthases and their regulators in human and mouse brown adipocytes in a YTHDF2/3-dependent manner. Collectively, these findings reveal a novel biological mechanism in the m6A-dependent regulation of the BAT secretome in modulating systemic insulin sensitivity in both mice and humans.

Presentation: 6/1/2024
8700 Mono(ADP-Ribosyl)ation of Ribosomal Proteins Promotes Differentiation During Adipogenesis
Authors: Tan X , Challa S , Camacho C V , Nandu T S , Stokes M S , Kraus W L
Published: October 2024

Disclosure: X. Tan: None. S. Challa: None. C.V. Camacho: None. T.S. Nandu: None. M.S. Stokes: None. W.L. Kraus: Other; Self; W.L.K. is a founder and stockholder for Ribon Therapeutics, Inc; a founder, member of the SAB, member of the BOD, and a stockholder for and ARase Therapeutics, Inc. He is also coholder of U.S. Patent.

ADP-ribosylation, a crucial post-translational modification, involves the covalent attachment of ADP-ribose units to amino acids on proteins through ADP-ribosyl transferase (ART) enzymes, utilizing NAD+ synthesized by nicotinamide mononucleotide adenylyl transferases (NMNATs). We previously showed that NMNAT-2, a cytoplasmic NAD+ synthase, increases during adipogenesis, leading to reduced nuclear NAD+ levels through compartmentalized NMN depletion and enhanced differentiation of preadipocytes. However, the implications of increased cytoplasmic NAD+ synthesis by NMNAT-2 in adipocytes remain unknown. Our current findings demonstrate elevated ribosome MARylation during 3T3-L1 preadipocyte differentiation, driven by NMNAT-2 induction. Inhibiting NAD+ biosynthesis by knockdown of NMNAT-2 or FK866 results in reduced ribosome MARylation, with PARP16 identified as the primary ART mediating MARylation. Knockdown of NMNAT-2 or PARP16 impaired global protein synthesis and preadipocyte differentiation. Treatment with the PARP16 inhibitor DB008 hindered ribosomal MARylation and preadipocyte differentiation. Immunofluorescence and subcellular fractionation analysis demonstrated that MARylated ribosomes are mostly associated with the endoplasmic reticulum. Using TMT mass spectrometry, we identified numerous high-confidence ADP-ribosylation sites on glutamate and aspartate residues in both undifferentiated and differentiated cells. We examined five proteins with specific sites of MARylation that are consistently more modified in the differentiated state. Among these, ADP-ribosylation of RPSA significantly promote preadipocyte differentiation; mutation of the site inhibits this effect. Ongoing investigations involve studying the impact of Parp16 knockout on high fat diet-induced obesity in mice to further determine the biological role of ribosome protein MARylation during adipogenesis.

This work is supported by a grant from the NIH/NIDDK (R01 DK069710) and the DOD OCRP (OC200311).

Presentation: 6/1/2024
7843 Nephron specific ATP6AP2 knockout reduces renal fat accumulation in mice consuming high fat diet
Authors: Allen Culver Silas , Lois Balugo Jamie Lynne , Jansen Maurits , Harris Thurl , Siragy Helmy M
Published: October 2024

Disclosure: S.A. Culver: None. J. Balugo: None. M. Jansen: None. T. Harris: None. H.M. Siragy: None.

Diet induced obesity causes increased renal fat accumulation in both humans and mice which contributes to obesity related kidney injury. We have previously shown that mice with nephron specific ATP6AP2 knockout (KO) consuming high fat diet (HFD) are resistant to weight gain due to increased urinary excretion of glucose, albumin, and fatty acids. Magnetic resonance imaging (MRI) is a useful imaging modality for quantifying organ fat content in live animals though there are few reports of its use quantifying fat in murine kidney. We hypothesized that nephron specific ATP6AP2 knockout inhibits renal fat accumulation with HFD and that we would be able to detect this using MRI. Eight-week-old male wild type (WT) or inducible nephron specific ATP6AP2 KO mice underwent either induction with oral doxycycline or received sucrose water as controls before being placed on either normal diet (ND) (12% fat) or HFD (45% fat) for 3 months. Mice were then imaged using a 9.4 Tesla Bruker BioSpin MRI to determine both liver and kidney cortex fat content. The mean weight of WT HFD mice was 36.03 g compared to 29.03 g in WT ND controls. ND fed and HFD fed KO mice weighed 22.74 g and 23.03 g respectively. MRI showed significant 67% (p<0.05) increase in liver fat content in WT HFD compared to WT ND fed controls, confirming the ability of our MRI technique to detect expected differences in organ fat content. In renal cortex, MRI detected a 54% (p<0.05) increase in lipid content for WT HFD mice compared to WT ND controls. Nephron specific ATP6AP2 KO on HFD demonstrated reduced renal cortical lipid content by 36% (p<0.05) compared to WT HFD controls. These results indicate that ATP6AP2 KO is associated with reduced renal fat accumulation that is measurable with in vivo MRI. MRI therefore represents a valuable tool for studying renal sequelae of obesity in live murine models. Future investigation should examine whether ATP6AP2 KO prevents renal lipotoxicity and kidney injury in the setting of obesity.

Presentation: 6/3/2024
6468 Once-Weekly Semaglutide and Taste Perception in Women with Obesity
Authors: Sever M Jensterle , Kovac J , Vovk A , Battelino S , Battelino T , Janez A
Published: October 2024

Disclosure: M. Jensterle Sever: None. J. Kovac: None. A. Vovk: None. S. Battelino: None. T. Battelino: None. A. Janez: None.

Objective: Semaglutide is a glucagon-like peptide-1 (GLP-1) analogue that causes weight loss by reducing appetite and lowering the desire for energy-dense, salty, and spicy foods, thereby reducing energy intake. We hypothesized that semaglutide would improve the taste sensitivity and modulate the tongue transcriptomic profile associated with gustatory coding in women with obesity.Design: We conducted a 16-week, single-blinded, randomized, placebo-controlled clinical trial involving 30 women aged 33.7 ± 6.1 years with a body mass index of 36.4 ± 4.4 kg/m2, randomized in a 1:1 ratio to once-weekly semaglutide 1.0 mg or a placebo (ClinicalTrials.gov number, NCT04263415). Methods: Taste sensitivity was assessed by strips impregnated with different concentrations of four basic tastes. Biopsies of the tongue were performed for the gene expression analysis. The brain responses were evaluated by functional MRI with the sweet solution dripping onto the tongue before and after a standardized meal intake. Results: Semaglutide improved the taste sensitivity expressed as a mean (± SD) from 11.9 ± 1.9 points to 14.4 ± 1.0 points, with an estimated treatment difference (ETD) of 2.5 points [95% CI, 1.7 to 3.3]. Genes EYA, PRMT8, CRLF1, and CYP1B1, associated with taste signaling transduction pathways, neural plasticity, and renewal of taste buds in the tongue, exhibited differential RNA expression across all tests performed by a multi-tiered analytical pipeline. Semaglutide increased functional activity in the angular gyrus of the parietal cortex in response to tasting the sweet solution compared to the distilled water 30 minutes after a standardized meal intake (semaglutide vs. placebo, p<0.001). Angular gyrus functions as a cross-modal hub where converging multisensory information is combined and integrated to comprehend and give sense to events and stimuli, manipulate mental representations, and solve familiar problems. It expresses GLP-1 receptor and is a part of the default mode network associated with shifting attention between rewarding and neutral stimuli, which has reduced connectivity in people with obesity. Conclusion: The present study provided evidence that semaglutide improved taste sensitivity, altered gene expression in the tongue linked to taste perception, and altered functional brain activity in an integrative hub beyond hypothalamic and midbrain reward circuitry in response to the sweet taste stimulus. Future studies will clarify whether the efficacy of semaglutide in treating obesity is also a »matter of taste«.

Presentation: 6/1/2024
8369 Overexpression of Neurosecretory Protein GL, a Novel Small Secretory Protein, Induces Fat Accumulation in Mice
Authors: Ukena Kazuyoshi , Narimatsu Yuki , Ogasawara Ayano , Furumitsu Megumi , Iwakoshi-Ukena Eiko
Published: October 2024

Disclosure: K. Ukena: None. Y. Narimatsu: None. A. Ogasawara: None. M. Furumitsu: None. E. Iwakoshi-Ukena: None.

Hypothalamic neuropeptides and peripheral factors influence food intake, body mass, and energy expenditure. For instance, neuropeptide Y (NPY)/agouti-related peptide (AgRP) are potent orexigenic factors and α-melanocyte-stimulating hormone (α-MSH) is an anorexigenic factor, respectively. In addition, leptin and insulin derived from peripheral tissues act on the hypothalamus to influence energy homeostasis. Precise hypothalamic control of energy balance affects puberty, thermoregulation, energy storage, and survival during different life-history stages. We have previously discovered a novel cDNA encoding the precursor of a neurosecretory small protein in the hypothalamus of birds and rodents (1, 2). The precursor protein contained a signal peptide sequence, a mature peptide sequence with 80 amino acid residues, a glycine amidation signal, and a dibasic amino acid cleavage site. Because the predicted C-terminal amino acids of the peptide were Gly-Leu-NH2, the small protein was named neurosecretory protein GL (NPGL). Quantitative RT-PCR analysis demonstrated that NPGL mRNA was expressed exclusively in the mediobasal hypothalamus (MBH), including the arcuate nucleus, and the expression was elevated on food deprivation in rodents. This finding led us to predict that NPGL was involved in the regulation of energy homeostasis. Indeed, using NPGL administration and neutralizing antibody administration in rats, we established that NPGL increased lipid accumulation in white adipose tissue (3). In the present study, to investigate the physiological function of NPGL in mice, we induced its overexpression using an adeno-associated virus (AAV) vector in NPGL-Cre driver mice. We then monitored food intake and recorded body mass changes. Npgl overexpression increased body mass. We also found that the overexpression resulted in increased masses of white adipose tissue and interscapular brown adipose tissue. Taken together, these results provide evidence for NPGL as a novel regulator of fat deposition in adipose tissues. Reference: (1) Ukena et al., Biochem. Biophys. Res. Commun. 2014 446:298-303. (2) Ukena, Gen. Comp. Endocrinol. 2018 260:164-170. (3) Iwakoshi-Ukena et al., eLife 2017 6:e28527.

Presentation: 6/3/2024
7215 Skeletal Muscle Mass Loss with Use of Dual GLP-1 GIP Receptor Agonist Tirzepitide
Authors: Zinn Jessica , Poretsky Leonid
Published: October 2024

Disclosure: J. Zinn: None. L. Poretsky: None.

Background: Multiple studies have demonstrated that treatment with tirzepitide results in significant improvement in HgbA1c as well as meaningful weight loss. Although the loss of skeletal muscle mass has been well described with the use of GLP-1 receptor agonists, there is little information regarding the effect of dual GLP-1/GIP receptor agonist tirzepitide on skeletal muscle mass. Clinical Case: A 68- year old male presented with a weight of 204.9 lbs and a BMI of 31.2 kg/m2. On exam, fat distribution was primarily central. Patient had a history of prediabetes, with an HgbA1c of 5.9%. He was unable to lose weight using lifestyle modifications, despite multiple meetings with a registered dietitian and consistent exercise of daily 3-6 mile walks. In the past, he used dulaglutide and semaglutide but had to discontinue these medications because of gastrointestinal side effects such as nausea and abdominal cramping. In June 2022, body composition (assessed using seca mBCA 514) demonstrated total weight of 204.9 lbs, BMI of 31.3 kg/m2, fat mass of 73.5 lbs (35.9%), fat-free mass of 131.4 lbs (64.1%), and skeletal muscle mass of 63.8 lbs (31.1%). He initiated Tirzepitide 2.5 mg weekly and maintained this dose for the duration of his treatment. Over nine months, total body weight decreased by 28.7 lbs or 13.9%. BMI decreased to 26.6 kg/m2 (at this point indicative of overweight status rather than obese). HgbA1c decreased to 5.3% (at this point no longer in the pre-diabetes range). Fat mass decreased by 18.9 lbs or 25.8%. Skeletal muscle mass decreased by 9.9 lbs or 15.5%. During this presentation we will provide a detailed time course of various metabolic and body composition parameters in this case and review relevant literature. Conclusion: We conclude that at least in some cases of patients treated with tirzepitide, up to 1/3 of lost body weight may be due to the loss of skeletal muscle mass. We suggest that, in addition to weight, serial body composition determinations should be obtained in patients on tirzepitide. It remains to be determined if adequate protein consumption and strength based exercise would ameliorate skeletal muscle mass loss observed with tirzepitide treatment. Reference: (1) Jastreboff AM, Aronne LJ, Ahmad NN, et al. Tirzepatide once weekly for the treatment of obesity. New England Journal of Medicine. 2022;387(3):205-216. doi:10.1056/nejmoa2206038

Presentation: 6/1/2024
8480 TIX100, a Novel Small Molecule for the Treatment of MASLD/MASH
Authors: Jo SeongHo , Chen Junqin , Jing Gu , Habegger Kirk M , Shalev Anath
Published: October 2024

Disclosure: S. Jo: None. J. Chen: None. G. Jing: None. K.M. Habegger: None. A. Shalev: Other; Self; TIXiMED, Inc. Founder & CSO.

Metabolic dysfunction-associated steatotic liver disease (MASLD) and steatohepatitis (MASH) (aka NAFLD and NASH) affect over 1 billion people worldwide, yet no approved therapy is currently available. MASLD/MASH is often associated with diabetes and obesity and some diabetes medications have shown some beneficial effects in this context. However, beyond steatosis these drugs are in general unable to prevent all the problems associated with MASH, such as inflammation, hepatocyte injury and especially fibrosis. Interestingly, we have now discovered that the new antidiabetic small molecule thioredoxin-interacting protein (TXNIP) inhibitor, TIX100, effectively protects against high-fat diet-induced hepatic steatosis. Oral TIX100 also protected obese and diabetic db/db mice against steatosis and significantly reduced the elevated expression levels of lipogenic genes found in db/db livers, many of which have been shown to be elevated in human MASLD. In addition though, TIX100 also normalized the blood glucose of db/db mice to the levels observed in lean control animals. Therefore, to address the question of whether TIX100 would still be effective in the absence of diabetes, obesity and any genetic leptin-receptor deficiency, we used a validated choline-deficient, amino acid-defined, high-fat diet to induce MASH. Indeed, compared to control chow, this diet significantly increased plasma liver enzymes indicating liver injury without causing diabetes or obesity. It also led to elevated hepatic expression of inflammatory and fibrogenic markers and histology revealed lipid accumulation and steatosis as well as pronounced fibrosis and upregulation of liver macrophages. Excitingly, all these pathological changes were blunted or improved by oral TIX100 treatment. Taken together, oral TIX100 may provide an attractive and effective novel approach for the treatment of MASLD/MASH that goes beyond the correction of diabetes, obesity, and steatosis.

Presentation: 6/3/2024
12411 Trends In Metabolic Dysfunction-associated Steatotic Liver Disease(MASLD) By Race/ethnicity: Data From NHANES 1988 To 2018
Authors: Shaheen M , Schrode K , Pan D , Najjar S M , Friedman T C
Published: October 2024

Disclosure: M. Shaheen: None. K. Schrode: None. D. Pan: None. S.M. Najjar: None. T.C. Friedman: None.

Societies of the liver recommended replacing the term non-alcoholic fatty liver disease (NAFLD) with metabolic dysfunction-associated steatotic liver disease (MASLD). This study aimed to investigate the trends in the MASLD prevalence by racial/ethnic among the U.S. adult population overtime. We analyzed data from 32,726 adults in National Health and Nutrition Examination Survey 1988-2018. Hepatic steatosis (HS) was diagnosed using the U.S. Fatty Liver Index. MASLD was diagnosed in individuals with both HS and at least one of the following: overweight/obesity, diabetes mellitus, hypertension, high triglyceride or HDL-Cholesterol or in treatment. Logistic regression was used to test for trends over time for the overall sample and for each racial/ethnic group accounting for design and sample weights. Prevalence estimates were age-adjusted using the direct standardization based on census data. We also estimated the percent change in the adjusted prevalence (change in the adjusted prevalence/baseline adjusted prevalence)*100. The study showed that over the study period, the age-adjusted prevalence of MASLD significantly increased from 16% in 1988 to 38% in 2018, representing a 138% percent increase. Concurrently, the age-adjusted prevalence of obesity increased from 23% in 1988 to 40% in 2018 reflecting a 74% increase (p<0.05). Among Mexican Americans, the age-adjusted MASLD prevalence consistently exceeded that of the overall population and other racial/ethnic groups. Significant increase in the percent change of the age-adjusted prevalence of MASLD was observed across all racial/ethnic groups (Mexican Americans=69%; Blacks=117%; Whites=140%) except Other Hispanics (no change). The age adjusted prevalence of obesity also increased significantly in all racial/ethnic groups (% change: Mexican Americans=58%; Other Hispanics=18%; Blacks=59%; Whites=77%) (p<0.05). Our findings indicated an increase in the age-adjusted prevalence of MASLD over time that surpassing the increase in obesity prevalence. These disparities warrant further investigation and targeted interventions to address MASLD effectively. Grants: R01MD012579, R24DA017298, U54MD007598, S21MD000103, UL1TR001881

Presentation: 6/2/2024