Distinct Molecular Mechanisms Research Articles

FLT4 causes developmental disorders of the cardiovascular and lymphovascular systems via pleiotropic molecular mechanisms.

Rare, deleterious genetic variants in FLT4 are associated with Tetralogy of Fallot (TOF), the most common cyanotic congenital heart disease (CHD). Distinct genetic variants in FLT4 are also an established cause of Milroy disease, the most prevalent form of primary hereditary lymphoedema. Phenotypic features of these two conditions are non-overlapping, implying pleiotropic cellular mechanisms during development. Here, we show that FLT4 variants identified in TOF patients, when expressed in primary human endothelial cells, cause aggregation of FLT4 protein in the perinuclear endoplasmic reticulum, activating proteostatic and metabolic signalling, whereas lymphoedema-associated FLT4 variants and wildtype FLT4 do not. FLT4 TOF variants display characteristic gene expression profiles in key developmental signalling pathways, revealing a role for FLT4 in cardiogenesis distinct from its role in lymphatic development. Inhibition of proteostatic signalling abrogates these effects, identifying potential avenues for therapeutic intervention. Depletion of flt4 in zebrafish caused cardiac phenotypes of reduced heart size and altered heart looping. These phenotypes were rescued with coinjection of wildtype human FLT4 mRNA, but incompletely or not at all by mRNA harbouring FLT4 TOF variants. Taken together, we identify a pathogenic mechanism for FLT4 variants predisposing to TOF that is distinct from the known dominant negative mechanism of Milroy-causative variants. FLT4 variants give rise to conditions of the two circulatory subdivisions of the vascular system via distinct developmental pleiotropic molecular mechanisms. Proteostatic dysfunction, if confirmed as a mechanism of CHD pathogenesis for other predisposing genes, may identify pathways to therapeutic interventions. Distinguishing mechanistically how variants in FLT4 give rise to CHD may have potential to individualise genetic counselling in affected families.

Impact of inflammatory preconditioning on murine microglial proteome response induced by focal ischemic brain injury.

Preconditioning with lipopolysaccharide (LPS) induces neuroprotection against subsequent cerebral ischemic injury, mainly involving innate immune pathways. Microglia are resident immune cells of the central nervous system (CNS) that respond early to danger signals through memory-like differential reprogramming. However, the cell-specific molecular mechanisms underlying preconditioning are not fully understood. To elucidate the distinct molecular mechanisms of preconditioning on microglia, we compared these cell-specific proteomic profiles in response to LPS preconditioning and without preconditioning and subsequent transient focal brain ischemia and reperfusion, - using an established mouse model of transient focal brain ischemia and reperfusion. A proteomic workflow, based on isolated microglia obtained from mouse brains by cell sorting and coupled to mass spectrometry for identification and quantification, was applied. Our data confirm that LPS preconditioning induces marked neuroprotection, as indicated by a significant reduction in brain infarct volume. The established brain cell separation method was suitable for obtaining an enriched microglial cell fraction for valid proteomic analysis. The results show a significant impact of LPS preconditioning on microglial proteome patterns by type I interferons, presumably driven by the interferon cluster regulator proteins signal transducer and activator of transcription1/2 (STAT1/2).

The molecular principles underlying diverse functions of the SLC26 family of proteins

Mammalian SLC26 proteins are membrane-based anion transporters that belong to the large SLC26/SulP family, and many of their variants are associated with hereditary diseases. Recent structural studies revealed a strikingly similar homodimeric molecular architecture for several SLC26 members, implying a shared molecular principle. Now a new question emerges as to how these structurally similar proteins execute diverse physiological functions. In this study we sought to identify the common vs. distinct molecular mechanism among the SLC26 proteins using both naturally occurring and artificial missense changes introduced to SLC26A4, SLC26A5, and SLC26A9. We found: (i) the basic residue at the anion binding site is essential for both anion antiport of SLC26A4 and motor functions of SLC26A5, and its conversion to a nonpolar residue is crucial but not sufficient for the fast uncoupled anion transport in SLC26A9; (ii) the conserved polar residues in the N- and C-terminal cytosolic domains are likely involved in dynamic hydrogen-bonding networks and are essential for anion antiport of SLC26A4 but not for motor (SLC26A5) and uncoupled anion transport (SLC26A9) functions; (iii) the hydrophobic interaction between each protomer’s last transmembrane helices, TM14, is not of functional significance in SLC26A9 but crucial for the functions of SLC26A4 and SLC26A5, likely contributing to optimally orient the axis of the relative movements of the core domain with respect to the gate domains within the cell membrane. These findings advance our understanding of the molecular mechanisms underlying the diverse physiological roles of the SLC26 family of proteins.

Potent Anthelmintic Activity of Chalcones Synthesized by an Effective Green Approach.

There is currently an urgent need for new anthelmintic agents due to increasing resistance to the limited available drugs. The chalcone scaffold is a privileged structure for developing new drugs and has been shown to exhibit potential antiparasitic properties. We synthesized a series of chalcones via Claisen-Schmidt condensation, introducing a novel recoverable catalyst derived from biochar obtained from the pyrolysis of tree pruning waste. Employing microwave irradiation and a green solvent, this approach demonstrated significantly reduced reaction times and excellent compatibility with various functional groups. The result was the generation of a library of functionalized chalcones, exhibiting exclusive (E)-selectivity and high to excellent yields. The chalcone derivatives were evaluated on the free-living nematode Caenorhabditis elegans. The chalcone scaffold, along with two derivatives incorporating a methoxy substituent in either ring, caused a concentration-dependent decrease of worm motility, revealing potent anthelmintic activity and spastic paralysis not mediated by the nematode levamisole-sensitive nicotinic receptor. The combination of both methoxy groups in the chalcone scaffold resulted in a less potent compound causing worm hypermotility at the short term, indicating a distinct molecular mechanism. Through the identification of promising drug candidates, this work addresses the demand for new anthelmintic drugs while promoting sustainable chemistry.

Hematopoietic stem cell niche generation and maintenance are distinguishable by an epitranscriptomic program.

The niche is typically considered as a pre-established structure sustaining stem cells. Therefore, the regulation of its formation remains largely unexplored. Whether distinct molecular mechanisms control the establishment versus maintenance of a stem cell niche is unknown. To address this, we compared perinatal and adult bone marrow mesenchymal stromal cells (MSCs), a key component of the hematopoietic stem cell (HSC) niche. MSCs exhibited enrichment in genes mediating m6A mRNA methylation at the perinatal stage and downregulated the expression of Mettl3, the m6A methyltransferase, shortly after birth. Deletion of Mettl3 from developing MSCs but not osteoblasts led to excessive osteogenic differentiation and a severe HSC niche formation defect, which was significantly rescued by deletion of Klf2, an m6A target. In contrast, deletion of Mettl3 from MSCs postnatally did not affect HSC niche. Stem cell niche generation and maintenance thus depend on divergent molecular mechanisms, which may be exploited for regenerative medicine.

Abstract 5651: Personalized transcriptomic profiling for advanced prostate cancer: Guiding second-generation hormonal drug selection

Abstract Introduction: Second-generation hormonal drugs have demonstrated significant efficacy in extending the survival of patients with advanced prostate cancer. These drugs operate through two distinct molecular mechanisms: inhibition of androgen biosynthesis and blockade of the androgen receptor. However, due to concerns of side effects, concurrent usage of multiple drugs is not recommended. A notable challenge is the eventual development of drug resistance, rendering all second-generation hormonal therapies ineffective. Currently, there is a lack of guidelines for helping patients choose the most suitable drug. In the context of pharmacogenomics, genetic differences can influence drug susceptibility and efficacy, with variations in genomic sequences and regulatory mechanisms impacting drug response at the transcriptional and translational levels. Considered the rapid processing capabilities of Affymetrix transcriptomic microarrays, we hypothesized that distinct gene signatures in patients who respond well or poorly to second-generation hormonal drugs could be identified and could act as a companion diagnostic tool. Methods: We enrolled 180 advanced prostate cancer patients, who fulfilled Taiwan national health insurance criteria of using second-generation hormonal drugs, from seven medical centers or regional hospitals in Taiwan at 3 time points: prior to drug administration, 3 months after treatment started and drug resistance developed. The RNA expression data and clinical parameters were analyzed, and classification models were built using IPA and R. Good or poor responders were defined by 3-month prostate-specific antigen (PSA) lowered by 90 percent. Results:: Differential expression and enrichment analyses post-treatment highlighted the activation of NF-kappa B signaling and various immune cell pathways. Notably, pathways related to primary immunodeficiency and IL-17 became prominent following the development of drug resistance. At the initial stages, distinct alterations in cell cycle pathways, particularly G1/S and G2/M, were observed between good and poor responders. This was elucidated through principal component analysis and subsequent dbscan clustering. To further distinguish between these responder groups, we utilized an integrative approach for feature selection, combining support vector machine, random forest classifier, and lasso regression. Despite the innovative approach, the model's error rate stood at approximately 15%, attributed mainly to the limited number of cases. Conclusion: Our study highlights the potential of using transcriptomic analysis to predict responses to second-generation hormonal drugs in advanced prostate cancer, paving the way for personalized treatment strategies. However, the need for larger cohort studies is evident to refine these predictive models and enhance their accuracy. Citation Format: Li-Hsin Chang, Hao-Han Chang, Xin-Yi Lin, Shu-Chi Wang, Chih-Pin Chuu, Deng-Neng Chen, Shu-Pin Huang, Chia-Yang Li. Personalized transcriptomic profiling for advanced prostate cancer: Guiding second-generation hormonal drug selection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5651.

Cellular Dynamics of Fas-Associated Death Domain in the Regulation of Cancer and Inflammation.

Fas-associated death domain (FADD) is an adaptor protein that predominantly transduces the apoptosis signal from the death receptor (DR) to activate caspases, leading to the initiation of apoptotic signaling and the coordinated removal of damaged, infected, or unwanted cells. In addition to its apoptotic functions, FADD is involved in signaling pathways related to autophagy, cell proliferation, necroptosis, and cellular senescence, indicating its versatile role in cell survival and proliferation. The subcellular localization and intracellular expression of FADD play a crucial role in determining its functional outcomes, thereby highlighting the importance of spatiotemporal mechanisms and regulation. Furthermore, FADD has emerged as a key regulator of inflammatory signaling, contributing to immune responses and cellular homeostasis. This review provides a comprehensive summary and analysis of the cellular dynamics of FADD in regulating programmed cell death and inflammation through distinct molecular mechanisms associated with various signaling pathways.

Pseudomonas aeruginosa MipA-MipB envelope proteins act as new sensors of polymyxins.

Due to the rising incidence of antibiotic-resistant infections, the last-line antibiotics, polymyxins, have resurged in the clinics in parallel with new bacterial strategies of escape. The Gram-negative opportunistic pathogen Pseudomonas aeruginosa develops resistance to colistin/polymyxin B by distinct molecular mechanisms, mostly through modification of the lipid A component of the LPS by proteins encoded within the arnBCDATEF-ugd (arn) operon. In this work, we characterized a polymyxin-induced operon named mipBA, present in P. aeruginosa strains devoid of the arn operon. We showed that mipBA is activated by the ParR/ParS two-component regulatory system in response to polymyxins. Structural modeling revealed that MipA folds as an outer-membrane β-barrel, harboring an internal negatively charged channel, able to host a polymyxin molecule, while the lipoprotein MipB adopts a β-lactamase fold with two additional C-terminal domains. Experimental work confirmed that MipA and MipB localize to the bacterial envelope, and they co-purify in vitro. Nano differential scanning fluorimetry showed that polymyxins stabilized MipA in a specific and dose-dependent manner. Mass spectrometry-based quantitative proteomics on P. aeruginosa membranes demonstrated that ∆mipBA synthesized fourfold less MexXY-OprA proteins in response to polymyxin B compared to the wild-type strain. The decrease was a direct consequence of impaired transcriptional activation of the mex operon operated by ParR/ParS. We propose MipA/MipB to act as membrane (co)sensors working in concert to activate ParS histidine kinase and help the bacterium to cope with polymyxin-mediated envelope stress through synthesis of the efflux pump, MexXY-OprA.IMPORTANCEDue to the emergence of multidrug-resistant isolates, antibiotic options may be limited to polymyxins to eradicate Gram-negative infections. Pseudomonas aeruginosa, a leading opportunistic pathogen, has the ability to develop resistance to these cationic lipopeptides by modifying its lipopolysaccharide through proteins encoded within the arn operon. Herein, we describe a sub-group of P. aeruginosa strains lacking the arn operon yet exhibiting adaptability to polymyxins. Exposition to sub-lethal polymyxin concentrations induced the expression and production of two envelope-associated proteins. Among those, MipA, an outer-membrane barrel, is able to specifically bind polymyxins with an affinity in the 10-µM range. Using membrane proteomics and phenotypic assays, we showed that MipA and MipB participate in the adaptive response to polymyxins via ParR/ParS regulatory signaling. We propose a new model wherein the MipA-MipB module functions as a novel polymyxin sensing mechanism.

Quantitative association between gene expression and blood cell production of individual hematopoietic stem cells in mice.

Individual hematopoietic stem cells (HSCs) produce different amounts of blood cells upon transplantation. Taking advantage of the intercellular variation, we developed an experimental and bioinformatic approach to evaluating the quantitative association between gene expression and blood cell production across individual HSCs. We found that most genes associated with blood production exhibit the association only at some levels of blood production. By mapping gene expression with blood production, we identified four distinct patterns of their quantitative association. Some genes consistently correlate with blood production over a range of levels or across all levels, and these genes are found to regulate lymphoid but not myeloid production. Other genes exhibit one or more clear peaks of association. Genes with overlapping peaks are found to be coexpressed in other tissues and share similar molecular functions and regulatory motifs. By dissecting intercellular variations, our findings revealed four quantitative association patterns that reflect distinct dose-response molecular mechanisms modulating the blood cell production of HSCs.

Heterozygous mutations in the C-terminal domain of COPA underlie a complex autoinflammatory syndrome.

Mutations in the N-terminal WD40 domain of coatomer protein complex subunit α (COPA) cause a type I interferonopathy, typically characterized by alveolar hemorrhage, arthritis, and nephritis. We described 3 heterozygous mutations in the C-terminal domain (CTD) of COPA (p.C1013S, p.R1058C, and p.R1142X) in 6 children from 3 unrelated families with a similar syndrome of autoinflammation and autoimmunity. We showed that these CTD COPA mutations disrupt the integrity and the function of coat protein complex I (COPI). In COPAR1142X and COPAR1058C fibroblasts, we demonstrated that COPI dysfunction causes both an anterograde ER-to-Golgi and a retrograde Golgi-to-ER trafficking defect. The disturbed intracellular trafficking resulted in a cGAS/STING-dependent upregulation of the type I IFN signaling in patients and patient-derived cell lines, albeit through a distinct molecular mechanism in comparison with mutations in the WD40 domain of COPA. We showed that CTD COPA mutations induce an activation of ER stress and NF-κB signaling in patient-derived primary cell lines. These results demonstrate the importance of the integrity of the CTD of COPA for COPI function and homeostatic intracellular trafficking, essential to ER homeostasis. CTD COPA mutations result in disease by increased ER stress, disturbed intracellular transport, and increased proinflammatory signaling.

Biomineralization in a cold environment: Insights from shield compositions and transcriptomics of polar sternaspids (Sternaspidae, Polychaeta)

The survival and physiological functions of polar marine organisms are impacted by global climate changes. Investigation of the adaptation mechanisms underlying biomineralization in polar organisms at low temperatures is important for understanding mineralized organismal sensitivity to climate change. Here, we performed electron probe analysis on the shields of Antarctic polychaete Sternaspis sendalli and Arctic polychaete Sternaspis buzhinskajae (Sternaspidae), and sequenced the transcriptomes of the tissues surrounding shields to examine biomineral characteristics and adaptive mechanisms in persistently cold environments. Compared to the temperate relative species, the relative abundance of iron, phosphorus, calcium, magnesium, nitrogen, sulfur and silicon in two polar sternaspid shields was similar to Sternaspis chinensis. However, the diversity and expression levels of biomineralization-related shell matrix proteins differed between the polar and temperate species, suggesting distinct molecular mechanisms underlying shield formation in cold environments. Tubulin and cyclophilin were upregulated compared to the temperate species. Furthermore, 42 positively selected genes were identified in Antarctic S. sendalli, with functions in cytoskeletal structure, DNA repair, immunity, transcription, translation, protein synthesis, and lipid metabolism. Highly expressed genes in both polar species were associated with cytoskeleton, macromolecular complexes and cellular component biosynthesis. Overall, this study reveals conserved elemental composition yet distinct biomineralization processes in the shields of polar sternaspids. The unique expression of biomineralization related genes and other cold-adaptation related genes provide molecular insights into biomineralization in cold marine environments.

Revitalizing mitochondrial function and empowering spinal cord recovery using copper-catalyzed Pdots

Spinal cord injury (SCI) inflicts devastating consequences for health. While drugs have been employed to alleviate SCI symptoms, their complex synthesis procedures and the need for specialized drug delivery raise concerns regarding administration. Herein, we utilize a green synthesis protocol to produce polymer quantum dots (Pdots) from polyvinylpyrrolidone precursors, empowered by the catalytic prowess of copper. These Copper-catalyzed Pdots (CPdots) not only offer enhanced improved fluorescence properties but also exhibit remarkable antioxidant, anti-inflammatory, and mitochondrial function repair effects. An intriguing finding emerged from our research as CPdots were observed to activate the AMPK/PGC-1α/Sirt3 pathway-a distinctive molecular mechanism that combats mitochondrial dysfunction, and mitigates calcium overload associated with acute SCI. Furthermore, the rapid metabolism and exceptional biocompatibility of our CPdots reinforce their safety and efficacy.

Functional and pharmacological analyses of visual habituation learning in larval zebrafish

Habituation allows animals to learn to ignore persistent but inconsequential stimuli. Despite being the most basic form of learning, a consensus model on the underlying mechanisms has yet to emerge. To probe relevant mechanisms, we took advantage of a visual habituation paradigm in larval zebrafish, where larvae reduce their reactions to abrupt global dimming (a dark flash). We used Ca2+ imaging during repeated dark flashes and identified 12 functional classes of neurons that differ based on their rate of adaptation, stimulus response shape, and anatomical location. While most classes of neurons depressed their responses to repeated stimuli, we identified populations that did not adapt or that potentiated their response. These neurons were distributed across brain areas, consistent with a distributed learning process. Using a small-molecule screening approach, we confirmed that habituation manifests from multiple distinct molecular mechanisms, and we have implicated molecular pathways in habituation, including melatonin, oestrogen, and GABA signalling. However, by combining anatomical analyses and pharmacological manipulations with Ca2+ imaging, we failed to identify a simple relationship between pharmacology, altered activity patterns, and habituation behaviour. Collectively, our work indicates that habituation occurs via a complex and distributed plasticity processes that cannot be captured by a simple model. Therefore, untangling the mechanisms of habituation will likely require dedicated approaches aimed at sub-component mechanisms underlying this multidimensional learning process.

Functional and pharmacological analyses of visual habituation learning in larval zebrafish.

Habituation allows animals to learn to ignore persistent but inconsequential stimuli. Despite being the most basic form of learning, a consensus model on the underlying mechanisms has yet to emerge. To probe relevant mechanisms, we took advantage of a visual habituation paradigm in larval zebrafish, where larvae reduce their reactions to abrupt global dimming (a dark flash). We used Ca2+ imaging during repeated dark flashes and identified 12 functional classes of neurons that differ based on their rate of adaptation, stimulus response shape, and anatomical location. While most classes of neurons depressed their responses to repeated stimuli, we identified populations that did not adapt or that potentiated their response. These neurons were distributed across brain areas, consistent with a distributed learning process. Using a small-molecule screening approach, we confirmed that habituation manifests from multiple distinct molecular mechanisms, and we have implicated molecular pathways in habituation, including melatonin, oestrogen, and GABA signalling. However, by combining anatomical analyses and pharmacological manipulations with Ca2+ imaging, we failed to identify a simple relationship between pharmacology, altered activity patterns, and habituation behaviour. Collectively, our work indicates that habituation occurs via a complex and distributed plasticity processes that cannot be captured by a simple model. Therefore, untangling the mechanisms of habituation will likely require dedicated approaches aimed at sub-component mechanisms underlying this multidimensional learning process.

Rett and Rett-related disorders: Common mechanisms for shared symptoms?

Rett syndrome is a neurodevelopmental disorder caused by loss-of-function mutations in the methyl-CpG binding protein-2 (MeCP2) gene that is characterized by epilepsy, intellectual disability, autistic features, speech deficits, and sleep and breathing abnormalities. Neurologically, patients with all three disorders display microcephaly, aberrant dendritic morphology, reduced spine density, and an imbalance of excitatory/inhibitory signaling. Loss-of-function mutations in the cyclin-dependent kinase-like 5 (CDKL5) and FOXG1 genes also cause similar behavioral and neurobiological defects and were referred to as congenital or variant Rett syndrome. The relatively recent realization that CDKL5 deficiency disorder (CDD), FOXG1 syndrome, and Rett syndrome are distinct neurodevelopmental disorders with some distinctive features have resulted in separate focus being placed on each disorder with the assumption that distinct molecular mechanisms underlie their pathogenesis. However, given that many of the core symptoms and neurological features are shared, it is likely that the disorders share some critical molecular underpinnings. This review discusses the possibility that deregulation of common molecules in neurons and astrocytes plays a central role in key behavioral and neurological abnormalities in all three disorders. These include KCC2, a chloride transporter, vGlut1, a vesicular glutamate transporter, GluD1, an orphan-glutamate receptor subunit, and PSD-95, a postsynaptic scaffolding protein. We propose that reduced expression or activity of KCC2, vGlut1, PSD-95, and AKT, along with increased expression of GluD1, is involved in the excitatory/inhibitory that represents a key aspect in all three disorders. In addition, astrocyte-derived brain-derived neurotrophic factor (BDNF), insulin-like growth factor 1 (IGF-1), and inflammatory cytokines likely affect the expression and functioning of these molecules resulting in disease-associated abnormalities.

A multi-omics dataset for the analysis of frontotemporal dementia genetic subtypes

Understanding the molecular mechanisms underlying frontotemporal dementia (FTD) is essential for the development of successful therapies. Systematic studies on human post-mortem brain tissue of patients with genetic subtypes of FTD are currently lacking. The Risk and Modyfing Factors of Frontotemporal Dementia (RiMod-FTD) consortium therefore has generated a multi-omics dataset for genetic subtypes of FTD to identify common and distinct molecular mechanisms disturbed in disease. Here, we present multi-omics datasets generated from the frontal lobe of post-mortem human brain tissue from patients with mutations in MAPT, GRN and C9orf72 and healthy controls. This data resource consists of four datasets generated with different technologies to capture the transcriptome by RNA-seq, small RNA-seq, CAGE-seq, and methylation profiling. We show concrete examples on how to use the resulting data and confirm current knowledge about FTD and identify new processes for further investigation. This extensive multi-omics dataset holds great value to reveal new research avenues for this devastating disease.

Distinct heat response molecular mechanisms emerge in cassava vasculature compared to leaf mesophyll tissue under high temperature stress

With growing concerns over global warming, cultivating heat-tolerant crops has become paramount to prepare for the anticipated warmer climate. Cassava (Manihot esculenta Crantz), a vital tropical crop, demonstrates exceptional growth and productivity under high-temperature (HT) conditions. Yet, studies elucidating HT resistance mechanisms in cassava, particularly within vascular tissues, are rare. We dissected the leaf mid-vein from leaf, and did the comparative transcriptome profiling between mid-vein and leaf to figure out the cassava vasculature HT resistance molecular mechanism. Anatomical microscopy revealed that cassava leaf veins predominantly consisted of vasculature. A thermal imaging analysis indicated that cassava experienced elevated temperatures, coinciding with a reduction in photosynthesis. Transcriptome sequencing produced clean reads in total of 89.17G. Using Venn enrichment, there were 65 differentially expressed genes (DEGs) and 93 DEGs had been found highly specifically expressed in leaf and mid-vein. Further investigation disclosed that leaves enhanced pyruvate synthesis as a strategy to withstand high temperatures, while mid-veins fortified themselves by bolstering lignin synthesis by comprehensive GO and KEGG analysis of DEGs. The identified genes in these metabolic pathways were corroborated through quantity PCR (QPCR), with results aligning with the transcriptomic data. To verify the expression localization of DEGs, we used in situ hybridization experiments to identify the expression of MeCCoAMT(caffeoyl-coenzyme A-3-O-methyltransferase) in the lignin synthesis pathway in cassava leaf veins xylem. These findings unravel the disparate thermotolerance mechanisms exhibited by cassava leaves and mid-veins, offering insights that could potentially inform strategies for enhancing thermotolerance in other crops.

Eltrombopag Plus Rhtpo for Corticosteroid-Resistant or Relapsed ITP: A Prospective Observed Study

During the COVID-19 pandemic, the classical subsequent treatment regimen for immune thrombocytopenia (ITP) of immunosuppressants and/or steroids might increase patients' susceptibility of virus infections. To minimize ITP patients' risk during the COVID-19 global crisis and to improve treatment efficacy, non-immunosuppressive therapies, mainly including thrombopoietin mimetics, such as eltrombopag or recombinant human thrombopoietin (rhTPO), may be better for them. Eltrombopag, a small molecule agonist of thrombopoietin receptor (TPO-RA), was recommended as the subsequent treatment for ITP patients, which also already showed robust efficacy. However, inefficacy, loss of response and fluctuation of platelet counts during treatment were the serious problems in eltrombopag treatment for corticosteroid-resistant or relapsed ITP.RhTPO is a full-length glycosylated-TPO produced by Chinese hamster ovary cells, which showed its effectiveness in ITP in a variety of studies.For the difference of rh-TPO and eltrombopag , it had distinct molecular structures, treatment mechanisms, and pharmacokinetic characteristics and thus different response patterns and side effect profile. Previous studies have confirmed the safety and effectiveness of switching between rh-TPO and eltrombopag for ITP patients who had no response to or experienced adverse events with their first TPO-RA.However, no study has focused on the safe and effectiveness of the combination therapy of eltrombopag and rhTPO.The investigators hypothesized that the combination of these two agents could be a promising option for ITP treatment. Methods This is a prospective observed study to investigate the efficacy and safety of eltrombopag plus rhTPO as treatment for corticosteroid-resistant or relapsed ITP during the COVID-19 pandemic. Participation Criteria The inclusion criteria were as follows:Clinically confirmed corticosteroid-resistant or relapsed primary ITP;Platelet count less than 30×10^9/L on two occasions or platelets above 30×10^9/L combined with bleeding manifestation (WHO bleeding scale 2 or above);≥ 18 years. Exclusion Criteria: with secondary ITP such as drug-related thrombocytopenia, viral infection-induced thrombocytopenia, or a known diagnosis of another autoimmune disease; with other possible causes of thrombocytopenia, such as leukaemia, myelodysplastic syndrome, and aplastic anaemia; impaired renal function as indicated by a serum creatinine level > 2.0 mg/dL;inadequate liver function as indicated by a total bilirubin level > 2.0 mg/dL and/or an aspartate aminotransaminase or alanine aminotransferase level > 3×upper limit of normal; with a New York Heart Classification III or IV heart disease; with a history of severe psychiatric disorder or are unable to comply with study and follow-up procedures; pregnant or lactating women, or plan to become pregnant or impregnated within 12 months of receiving study drug; previous splenectomy; with previous or concomitant malignant disease; expected survival of < 2 years;not willing to participate in the study. Procedure For the patients with the combination therapy of eltrombopag and rhTPO, they received eltrombopag 25-75 mg oral daily and rh-TPO 300U/kg subcutaneous injection once daily for 7 consecutive days, followed by a tapering dose in maintenance therapy to maintain the platelet count between 50-150×10 9/L. Results From September 2nd, 2020, to July 1st, 2023, a total of 29 patients with corticosteroid-resistant or relapsed ITP were enrolled to receive eltrombopag plus rhTPO treatment, which included 11 male and 18 female patients.The median age were 55 (23-91) years old. The baseline platelete count was 7 (1-17)×10 9/L. Most patients (69.0%) were with severe ITP (platelet counts≤10×10 9/L or bleeding score ≥5).Response was defined as the achievement of platelet counts ≥30 × 10 9/L and ≥2 times the baseline platelet count at least once with treatment.The response rates at 1 week (early response) and 1 month ( initial response) were 37.9%(11/29) and 62.1%(18/29), respectively. After 6 weeks' treatment, 82.8% (25/29) of patients achieved response, and the improvement of bleeding symptoms were observed in most patients (89.7%).No thromboembolic events occurred during treatment. Conclusion Elrtombopag and rhTPO was a safe and effective therapy for corticosteroid-resistant or relapsed ITP, especially for severe ITP.

Distinct neurogenetic mechanisms establish the same chemosensory valence state at different life stages in Caenorhabditis elegans.

An animal's preference for many chemosensory cues remains constant despite dramatic changes in the animal's internal state. The mechanisms that maintain chemosensory preference across different physiological contexts remain poorly understood. We previously showed that distinct patterns of neural activity and motor output are evoked by carbon dioxide (CO2) in starved adults vs dauers of Caenorhabditis elegans, despite the two life stages displaying the same preference (attraction) for CO2. However, how the distinct CO2-evoked neural dynamics and motor patterns contribute to CO2 attraction at the two life stages remained unclear. Here, using a CO2 chemotaxis assay, we show that different interneurons are employed to drive CO2 attraction at the two life stages. We also investigate the molecular mechanisms that mediate CO2 attraction in dauers vs adults. We show that insulin signaling promotes CO2 attraction in dauers but not starved adults and that different combinations of neurotransmitters and neuropeptides are used for CO2 attraction at the two life stages. Our findings provide new insight into the distinct molecular and cellular mechanisms used by C. elegans at two different life stages to generate attractive behavioral responses to CO2.

Mitochondrial aminoacyl-tRNA synthetases trigger unique compensatory mechanisms in neurons.

Mitochondrial aminoacyl-tRNA synthetase (mt-ARS) mutations cause severe, progressive, and often lethal diseases with highly heterogeneous and tissue-specific clinical manifestations. This study investigates the molecular mechanisms triggered by three different mt-ARS defects caused by biallelic mutations in AARS2, EARS2, and RARS2, using an in vitro model of human neuronal cells. We report distinct molecular mechanisms of mitochondrial dysfunction among the mt-ARS defects studied. Our findings highlight the ability of proliferating neuronal progenitor cells (iNPCs) to compensate for mitochondrial translation defects and maintain balanced levels of oxidative phosphorylation (OXPHOS) components, which becomes more challenging in mature neurons. Mutant iNPCs exhibit unique compensatory mechanisms, involving specific branches of the integrated stress response, which may be gene-specific or related to the severity of the mitochondrial translation defect. RNA sequencing revealed distinct transcriptomic profiles showing dysregulation of neuronal differentiation and protein translation. This study provides valuable insights into the tissue-specific compensatory mechanisms potentially underlying the phenotypes of patients with mt-ARS defects. Our novel in vitro model may more accurately represent the neurological presentation of patients and offer an improved platform for future investigations and therapeutic development.