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Advanced Science (Weinheim,... Jun 2024Cancer cells must develop strategies to adapt to the dynamically changing stresses caused by intrinsic or extrinsic processes, or therapeutic agents. Metabolic...
Cancer cells must develop strategies to adapt to the dynamically changing stresses caused by intrinsic or extrinsic processes, or therapeutic agents. Metabolic adaptability is crucial to mitigate such challenges. Considering metabolism as a central node of adaptability, it is focused on an energy sensor, the AMP-activated protein kinase (AMPK). In a subtype of pancreatic ductal adenocarcinoma (PDAC) elevated AMPK expression and phosphorylation is identified. Using drug repurposing that combined screening experiments and chemoproteomic affinity profiling, it is identified and characterized PF-3758309, initially developed as an inhibitor of PAK4, as an AMPK inhibitor. PF-3758309 shows activity in pre-clinical PDAC models, including primary patient-derived organoids. Genetic loss-of-function experiments showed that AMPK limits the induction of ferroptosis, and consequently, PF-3758309 treatment restores the sensitivity toward ferroptosis inducers. The work established a chemical scaffold for the development of specific AMPK-targeting compounds and deciphered the framework for the development of AMPK inhibitor-based combination therapies tailored for PDAC.
PubMed: 38885414
DOI: 10.1002/advs.202307695 -
Organic phosphate but not inorganic phosphate regulates expression through MAPK and TGF-ꞵ signaling.IScience Jun 2024One of the main regulators of phosphate homeostasis is fibroblast growth factor 23 (FGF23), secreted by osteocytes. The effects of organic versus inorganic dietary...
One of the main regulators of phosphate homeostasis is fibroblast growth factor 23 (FGF23), secreted by osteocytes. The effects of organic versus inorganic dietary phosphate on this homeostasis are unclear. This study used MC3T3-E1 FGF23-producing cells to examine the transcriptomic responses to these phosphates. Most importantly, the expression and secretion of FGF23 were only increased in response to organic phosphate. Gene ontology terms related to a response to environmental change were only enriched in cells treated with organic phosphate while cells treated with inorganic phosphate were enriched for terms associated with regulation of cellular phosphate metabolism. Inhibition of MAPK signaling diminished the response of to organic phosphate, suggesting it activates FGF23. TGF-β signaling inhibition increased expression after the addition of organic phosphate, while the negative TGF-β regulator decreased this response. In summary, the observed differential response of FGF23-producing to phosphate types may have consequences for phosphate homeostasis.
PubMed: 38883842
DOI: 10.1016/j.isci.2024.109625 -
Planta Jun 2024We generated transplastomic tobacco lines that stably express a human Basic Fibroblast Growth Factor (hFGFb) in their chloroplasts stroma and purified a biologically...
We generated transplastomic tobacco lines that stably express a human Basic Fibroblast Growth Factor (hFGFb) in their chloroplasts stroma and purified a biologically active recombinant hFGFb. MAIN: The use of plants as biofactories presents as an attractive technology with the potential to efficiently produce high-value human recombinant proteins in a cost-effective manner. Plastid genome transformation stands out for its possibility to accumulate recombinant proteins at elevated levels. Of particular interest are recombinant growth factors, given their applications in animal cell culture and regenerative medicine. In this study, we produced recombinant human Fibroblast Growth Factor (rhFGFb), a crucial protein required for animal cell culture, in tobacco chloroplasts. We successfully generated two independent transplastomic lines that are homoplasmic and accumulate rhFGFb in their leaves. Furthermore, the produced rhFGFb demonstrated its biological activity by inducing proliferation in HEK293T cell lines. These results collectively underscore plastid genome transformation as a promising plant-based bioreactor for rhFGFb production.
Topics: Nicotiana; Humans; Plants, Genetically Modified; Fibroblast Growth Factor 2; Chloroplasts; Recombinant Proteins; HEK293 Cells; Cell Proliferation; Plant Leaves
PubMed: 38878167
DOI: 10.1007/s00425-024-04456-5 -
ACS Nano Jul 2024The cellular environment, characterized by its intricate composition and spatial organization, hosts a variety of organelles, ranging from membrane-bound ones to...
The cellular environment, characterized by its intricate composition and spatial organization, hosts a variety of organelles, ranging from membrane-bound ones to membraneless structures that are formed through liquid-liquid phase separation. Cells show precise control over the position of such condensates. We demonstrate that organelle movement in external concentration gradients, , is distinct from the one of colloids because fluxes can remain finite inside the liquid-phase droplets and movement of the latter arises from incompressibility. Within cellular domains diffusiophoresis naturally arises from biochemical reactions that are driven by a chemical fuel and produce waste. Simulations and analytical arguments within a minimal model of reaction-driven phase separation reveal that the directed movement stems from two contributions: Fuel and waste are refilled or extracted at the boundary, resulting in concentration gradients, which (i) induce product fluxes via incompressibility and (ii) result in an asymmetric forward reaction in the droplet's surroundings (as well as asymmetric backward reaction inside the droplet), thereby shifting the droplet's position. We show that the former contribution dominates and sets the direction of the movement, toward or away from fuel source and waste sink, depending on the product molecules' affinity toward fuel and waste, respectively. The mechanism thus provides a simple means to organize condensates with different composition. Particle-based simulations and systems with more complex reaction cycles corroborate the robustness and universality of this mechanism.
PubMed: 38875706
DOI: 10.1021/acsnano.3c12842 -
Journal of Translational Medicine Jun 2024
Correction: Real‑world data suggest effectiveness of the allogeneic mesenchymal stromal cells preparation MSC‑FFM in ruxolitinib‑refractory acute graft‑versus‑host disease.
PubMed: 38872104
DOI: 10.1186/s12967-024-05348-8 -
Nature Communications Jun 2024Generating 3D bone cell networks in vitro that mimic the dynamic process during early bone formation remains challenging. Here, we report a synthetic biodegradable...
Generating 3D bone cell networks in vitro that mimic the dynamic process during early bone formation remains challenging. Here, we report a synthetic biodegradable microporous hydrogel for efficient formation of 3D networks from human primary cells, analysis of cell-secreted extracellular matrix (ECM) and microfluidic integration. Using polymerization-induced phase separation, we demonstrate dynamic in situ formation of microporosity (5-20 µm) within matrix metalloproteinase-degradable polyethylene glycol hydrogels in the presence of living cells. Pore formation is triggered by thiol-Michael-addition crosslinking of a viscous precursor solution supplemented with hyaluronic acid and dextran. The resulting microporous architecture can be fine-tuned by adjusting the concentration and molecular weight of dextran. After encapsulation in microporous hydrogels, human mesenchymal stromal cells and osteoblasts spread rapidly and form 3D networks within 24 hours. We demonstrate that matrix degradability controls cell-matrix remodeling, osteogenic differentiation, and deposition of ECM proteins such as collagen. Finally, we report microfluidic integration and proof-of-concept osteogenic differentiation of 3D cell networks under perfusion on chip. Altogether, this work introduces a synthetic microporous hydrogel to efficiently differentiate 3D human bone cell networks, facilitating future in vitro studies on early bone development.
Topics: Humans; Hydrogels; Mesenchymal Stem Cells; Osteogenesis; Cell Differentiation; Osteoblasts; Extracellular Matrix; Porosity; Cell Culture Techniques, Three Dimensional; Polyethylene Glycols; Tissue Engineering; Hyaluronic Acid; Cells, Cultured; Tissue Scaffolds; Dextrans
PubMed: 38871693
DOI: 10.1038/s41467-024-49280-3 -
Biophysical Journal Jun 2024Exchange of material across two membranes, as in the case of synaptic neurotransmitter release from a vesicle, involves the formation and poration of a hemifusion...
Exchange of material across two membranes, as in the case of synaptic neurotransmitter release from a vesicle, involves the formation and poration of a hemifusion diaphragm (HD). The nontrivial geometry of the HD leads to environment-dependent control, regarding the stability and dynamics of the pores required for this kind of exocytosis. This work combines particle simulations, field-based calculations, and phenomenological modeling to explore the factors influencing the stability, dynamics, and possible control mechanisms of pores in HDs. We find that pores preferentially form at the HD rim, and that their stability is sensitive to a number of factors, including the three line tensions, membrane tension, HD size, and the ability of lipids to "flip-flop" across leaflets. Along with a detailed analysis of these factors, we discuss ways that vesicles or cells may use them to open and close pores and thereby quickly and efficiently transport material.
PubMed: 38867448
DOI: 10.1016/j.bpj.2024.06.009 -
Biomacromolecules Jun 2024After RNAi was first discovered over 20 years ago, siRNA-based therapeutics are finally becoming reality. However, the delivery of siRNA has remained a challenge. In our...
After RNAi was first discovered over 20 years ago, siRNA-based therapeutics are finally becoming reality. However, the delivery of siRNA has remained a challenge. In our previous research, we found that spermine-based poly(β-amino ester)s are very promising for siRNA delivery. However, the role of hydrophobic modification in siRNA delivery of spermine-based poly(β-amino ester)s is not fully understood yet. In the current work, we synthesized spermine-based poly(β-amino ester)s with different percentages of oleylamine side chains, named P(SpOABAE). The chemical structures of the polymers were characterized by H NMR. The polymers showed efficient siRNA encapsulation determined by SYBR Gold assays. The hydrodynamic diameters of the P(SpOABAE) polyplexes from charge ratio N/P 1 to 20 were 30-100 nm except for aggregation phenomena observed at N/P 3. Morphology of the polyplexes was visualized by atomic force microscopy, and cellular uptake was determined by flow cytometry in H1299 cells, where all the polyplexes showed significantly higher cellular uptake than hyperbranched polyethylenimine (25 kDa). The most hydrophobic P(SpOABAE) polyplexes were able to achieve more than 90% GFP knockdown in H1299/eGFP cells. The fact that gene silencing efficacy increased with hydrophobicity but cellular uptake was affected by both charge and hydrophobic interactions highlights the importance of endosomal escape. For pulmonary administration and improved storage stability, the polyplexes were spray-dried. Results confirmed the maintained siRNA activity after storage for 3 months at room temperature, indicating potential for dry powder inhalation.
PubMed: 38866384
DOI: 10.1021/acs.biomac.4c00283 -
ACS Applied Materials & Interfaces Jun 2024Solid-state polymer electrolytes (SPEs), such as poly(ethylene oxide) (PEO), have good flexibility when compared to ceramic-type solid electrolytes. Therefore, it could...
Solid-state polymer electrolytes (SPEs), such as poly(ethylene oxide) (PEO), have good flexibility when compared to ceramic-type solid electrolytes. Therefore, it could be an ideal solid electrolyte for zero-excess all-solid-state Li metal battery (ZESSLB), also known as anode-free all-solid-state Li battery, development by offering better contact to the Cu current collector. However, the low Coulombic efficiencies observed from polymer type solid-state Li batteries (SSLBs) raise the concern that PEO may consume the limited amount of Li in ZESSLB to fail the system. Here, we designed ZESSLBs by using all-ceramic half-cells and an extra PEO electrolyte interlayer to study the reactivity between PEO and freshly deposited Li under a real battery operating conduction. By shuttling active Li back from the anode to the cathode, the PEO SPEs can be separated from the ZESSLBs for experimental studies without the influence from cathode materials or possible contamination from the usage of Li foil as the anode. Electrochemical cycling of ZESSLBs shows that the capacities of ZESSLBs with solvent-free and solvent-casted PEO SPEs significantly degraded compared to the ones with Li metal as the anode for the all-solid-state Li batteries. The fast capacity degradation of ZESSLBs using different types of PEO SPEs is evidenced to be associated with Li reacting with PEO, residual solvent, and water in PEO and dead Li formation upon the presence or absence of residual solvent. The results suggest that avoiding direct contact between the PEO electrolyte and deposited lithium is necessary when there is only a limited amount of Li available in ZESSLBs.
PubMed: 38863333
DOI: 10.1021/acsami.4c03387 -
Cell Death Discovery Jun 2024A key feature of cancer is the disruption of cell cycle regulation, which is characterized by the selective and abnormal activation of cyclin-dependent kinases (CDKs)....
A key feature of cancer is the disruption of cell cycle regulation, which is characterized by the selective and abnormal activation of cyclin-dependent kinases (CDKs). Consequently, targeting CDKs via meriolins represents an attractive therapeutic approach for cancer therapy. Meriolins represent a semisynthetic compound class derived from meridianins and variolins with a known CDK inhibitory potential. Here, we analyzed the two novel derivatives meriolin 16 and meriolin 36 in comparison to other potent CDK inhibitors and could show that they displayed a high cytotoxic potential in different lymphoma and leukemia cell lines as well as in primary patient-derived lymphoma and leukemia cells. In a kinome screen, we showed that meriolin 16 and 36 prevalently inhibited most of the CDKs (such as CDK1, 2, 3, 5, 7, 8, 9, 12, 13, 16, 17, 18, 19, 20). In drug-to-target modeling studies, we predicted a common binding mode of meriolin 16 and 36 to the ATP-pocket of CDK2 and an additional flipped binding for meriolin 36. We could show that cell cycle progression and proliferation were blocked by abolishing phosphorylation of retinoblastoma protein (a major target of CDK2) at Ser612 and Thr82. Moreover, meriolin 16 prevented the CDK9-mediated phosphorylation of RNA polymerase II at Ser2 which is crucial for transcription initiation. This renders both meriolin derivatives as valuable anticancer drugs as they target three different Achilles' heels of the tumor: (1) inhibition of cell cycle progression and proliferation, (2) prevention of transcription, and (3) induction of cell death.
PubMed: 38862521
DOI: 10.1038/s41420-024-02056-6