Macrophages modulate stiffness-related foreign body responses through plasma membrane deformation.

Implants are widely used in medical applications and yet macrophage-mediated foreign body reactions caused by implants severely impact their therapeutic effects. Although the extensive use of multiple surface modifications has been introduced to provide some mitigation of fibrosis, little is known about how macrophages recognize the stiffness of the implant and thus influence cell behaviors. Here, we demonstrated that macrophage stiffness sensing leads to differential inflammatory activation, resulting in different degrees of fibrosis. The potential mechanism for macrophage stiffness sensing in the early adhesion stages tends to involve cell membrane deformations on substrates with different stiffnesses. Combining theory and experiments, we show that macrophages exert traction stress on the substrate through adhesion and altered membrane curvature, leading to the uneven distribution of the curvature-sensing protein Baiap2, resulting in cytoskeleton remodeling and inflammation inhibition. This study introduces a physical model feedback mechanism for early cellular stiffness sensing based on cell membrane deformation, offering perspectives for future material design and targeted therapies.

Molecular and clinical characterization of two unrelated families with factor V deficiency, including a novel nonsense variant (p.Gln1532*).

BACKGROUND: Factor V (FV) is an essential cofactor in the coagulation cascade. The characterization of novel mutations is advantageous for the clinical management of FV-deficient patients. METHODS: Coagulation screening and thrombin generation assay were performed with the plate-poor plasma. All 25 exons of the F5 gene were amplified and sequenced. The ClustalX-2.1 software was applied to the multiple sequence alignment. The possible adverse effects of mutations were investigated with online bioinformatics software and protein modeling. RESULTS: Two unrelated families with FV deficiency were under investigation. Proband A was an 18-year-old youth with recurrent epistaxis. Proband B was a 29-year-old woman who did not present with any bleeding symptoms. Three heterozygous mutations (p.Gln1532*, p.Phe218Ser, and p.Asp2222Gly) were detected. Interestingly, they were compound heterozygotes and both contained the p.Asp2222Gly, a polymorphism. The thrombin generation assay showed that both patients had impaired ability of thrombin generation, and in particular, proband A was more severe. Conservation, pathogenicity and protein modeling studies all indicated that these three mutations could cause deleterious effects on the function and structure of FV. CONCLUSION: These three mutations are responsible for the FV-deficient in two pedigrees. Moreover, the nonsense variant p.Gln1532* is first reported in the world.

H2O2 Self-Supplying and GSH-Depleting Nanocatalyst for Copper Metabolism-Based Synergistic Chemodynamic Therapy and Chemotherapy.

Chemodynamic therapy (CDT) that involves the use of Fenton catalysts to convert endogenous hydrogen peroxide (H2O2) to hydroxyl radicals (·OH) constitutes a promising strategy for cancer therapy; however, insufficient endogenous H2O2 and glutathione (GSH) overexpression render its efficiency unsatisfactory. Herein, we present an intelligent nanocatalyst that comprises copper peroxide nanodots and DOX-loaded mesoporous silica nanoparticles (MSNs) (DOX@MSN@CuO2) and can self-supply exogenous H2O2 and respond to specific tumor microenvironments (TME). Following endocytosis into tumor cells, DOX@MSN@CuO2 initially decomposes into Cu2+ and exogenous H2O2 in the weakly acidic TME. Subsequently, Cu2+ reacts with high GSH concentrations, thereby inducing GSH depletion and reducing Cu2+ to Cu+ Next, the generated Cu+ undergoes Fenton-like reactions with exogenous H2O2 to accelerate toxic ·OH production, which exhibits a rapid reaction rate and is responsible for tumor cell apoptosis, thereby enhancing CDT. Furthermore, the successful delivery of DOX from the MSNs achieves chemotherapy and CDT integration. Thus, this excellent strategy can resolve the problem of insufficient CDT efficacy due to limited H2O2 and GSH overexpression. Integrating H2O2 self-supply and GSH deletion enhances CDT, and DOX-induced chemotherapy endows DOX@MSN@CuO2 with effective tumor growth-inhibiting properties alongside minimal side effects in vivo.

Gene Variants in Two Families with Inherited Coagulation Factor XI Deficiency and Identification of Mutations.

INTRODUCTION: Mutations in the F11 gene can cause factor XI (FXI) deficiency, leading to abnormal coagulation activity and injury-related bleeding tendency. Therefore, identifying F11 gene mutations and studying the molecular basis will help us understand the pathogenesis of FXI deficiency. METHODS: Coagulation tests and gene sequencing analysis of all members were performed. FXI wild-type and mutant expression plasmids were constructed and transfected into HEK293FT cells. The FXI protein expression level was evaluated by ELISA and Western blot. RESULTS: The FXI activity (FXI:C) and FXI antigen (FXI:Ag) of proband-1 were decreased to 2% and 5%, respectively. FXI:C and FXI:Ag of proband-2 were reduced to 15% and 32%, respectively. Four mutations were found in the two unrelated families, including c.536C>T (p.T179M), c.1556G>A (p.W519*), c.434A>G (p.H145R), and c.1325_1325delT (p.L442Cfs*8). In vitro studies in transiently transfected HEK293FT cells demonstrated that p.T179M, p.W519*, and p.L442Cfs*8 mutations significantly lowered the FXI levels in the culture media. The FXI levels in the culture media and cell lysates of p.H145R mutation were similar to the wild type. CONCLUSION: Our results confirm that the four mutations in the F11 gene are causative in the 2 FXI deficiency families. Moreover, the p.H145R mutation is a cross-reactive material (CRM)-positive phenotype. The other three mutations are CRM-negative phenotypes and lead to FXI protein secretion disorder.

Subcellular-Targeted Near-Infrared-Responsive Nanomedicine with Synergistic Chemo-photothermal Therapy against Multidrug Resistant Cancer.

Multidrug resistance (MDR) is a major obstacle to effective cancer treatment. Therefore, developing effective approaches for overcoming the limitation of MDR in cancer therapy is very essential. Chemotherapy combined with photothermal therapy (PTT) is a potential therapeutic option against MDR. Herein, we developed a subcellular-targeted near-infrared (NIR)-responsive nanomedicine (Fe3O4@PDA-TPP/S2-PEG-hyd-DOX, abbreviated as Fe3O4-ATSPD) as a new photothermal agent with improved photothermal stability and efficiency. This system demonstrates high stability in blood circulation and can be accumulated at the tumor site by magnetic targeting enhanced permeability and retention effect (EPR). Near-infrared (NIR) irradiation at the tumor site generates a photothermal effect from the photosensitizer Fe3O4@PDA, leading to a dramatic decrease in mitochondrial membrane potential. Simultaneously, the conjugated drugs released under low pH condition in endosomes or lysosomes cause nucleus DNA damage and cell apoptosis. This subcellular-targeted NIR-responsive nanomedicine with efficient integration of diagnosis and therapy could significantly enhance MDR cancer treatment by combination of chemotherapy and PTT.

Analysis of phenotype and gene mutation in three pedigrees with inherited antithrombin deficiency.

BACKGROUND: Inherited AT deficiency is an autosomal-dominant thrombophilic disorder usually caused by various SERPINC1 defects associated with a high risk of recurrent venous thromboembolism. In this article, the phenotype, gene mutation, and molecular pathogenic mechanisms were determined in three pedigrees with inherited AT deficiency. METHODS: Coagulation indices were examined on STAGO STA-R-MAX analyzer. The AT:Ag was analyzed by ELISA. All exons and flanking sequences of SERPINC1 were amplified by PCR. AT wild type and three mutant expression plasmids were constructed and then transfected into HEK293FT cells. The expression level of AT protein was analyzed by ELISA and Western blot. RESULTS: The AT:A and AT:Ag of probands 1 and 3 were decreased to 49% and 52 mg/dL, 38% and 44 mg/dL, respectively. The AT:A of proband 2 was decreased to 32%. The SERPINC1 gene analysis indicated that there was a p.Ile421Thr in proband 1, a p.Leu417Gln in proband 2, and a p.Met252Thr in proband 3, respectively. The AT mRNA expression level of the three mutants was not significantly different from AT-WT by qRT-PCR. The results of ELISA and Western blot tests showed that the AT-M252T and AT-I421T mutants had a higher AT expression than the AT wild type (AT-WT), and the AT protein expression of AT-L417Q mutants had no significant difference compared with AT-WT in the cell lysate. The AT expression levels of AT-M252T and AT-I421T mutants were lower than that of AT-WT, and there was no significant difference between AT-L417Q mutant and AT-WT in the supernatant. CONCLUSION: The p.I421T and p.M252T mutations affected the secretion of AT protein leading to type I AT deficiency of probands 1 and 3. The p.Leu417Gln mutation was responsible for the impaired or ineffective activity AT protein in proband 2 and caused type II AT deficiency.

[Analysis of the molecular pathogenesis of hereditary protein C deficiency due to a p.Gly86Asp variant of the PROC gene].

OBJECTIVE: To explore the molecular pathogenesis of hereditary protein C (PC) deficiency due to a p.Gly86Asp variant of the PROC gene through in vitro expression experiment. METHODS: Wild type and Gly86Asp mutant expression plasmids of PC were constructed and respectively transfected into HEK 293FT cells. Total RNA was extracted from the transfected cells, and the expression of PROC gene was determined by quantitative real-time PCR (qRT-PCR). PC antigen (PC:Ag) in the supernatant of cell culture and cell lysate was determined by enzyme-linked immunosorbent assay (ELISA), and the level of PC protein was detected by Western blotting. RESULTS: qRT-PCR has detected no significant difference in the transcription level of wild-type and mutant-type PC. Compared with the wild type, the level of mutant PC:Ag in the supernatant and cell lysate were 81.3%±2.6% and 110.0%±2.8%, respectively. No difference was detected in the molecular weight between the wild-type and mutant-type PC by Western blotting. The PC content of mutant type was higher than wild-type in cell lysate, while the opposite was found with the cell culture supernatant. CONCLUSION: The impaired secretion by mutant PC may be the molecular mechanism of PC deficiency caused by the p.Gly86Asp variant.

[Analysis of genetic variant in a Chinese pedigree with hereditary factor VIII deficiency].

OBJECTIVE: To explore the genetic basis for a patient with factor VIII deficiency. METHODS: All exons of the F13A1 and F13B genes were amplified by PCR and sequenced directly. The sequencing was performed with a reverse primer if a variant was found. Conservation of variant site was analyzed by the ClustalX software. Four online bioinformatic software including Mutation Taster, PolyPhen-2, PROVEAN and SIFT were used to predict the function of the mutation site. The Swiss-PdbViewer software was applied to analyze the changes in the protein model and intermolecular force. RESULTS: The proband was found to harbor a novel c.515G>C (p.Arg171Pro) variant of the F13A1 gene. The corresponding amino acid Arg171 is conserved among homologous species. Bioinformatic analysis indicated that Arg171Pro variant may affect the protein function. Protein model analysis showed that in the wild-type, there is one hydrogen bond between Arg171 and Pro27; one hydrogen bond between Arg171 and Thr28; two hydrogen bonds between Arg171 and Glu102. When Arg171 was mutated to Pro171, the three hydrogen bonds between Arg171 and Pro27, Glu102 are all disappeared and formed a new benzene ring which might affect the stability of the protein structure. No variant was found in the F13B gene. CONCLUSION: The Arg171Pro variant may account for the decreased FVIII level. Above finding has enriched the spectrum of F13A1 gene variants.

Metallo-Helicoid with Double Rims: Polymerization Followed by Folding by Intramolecular Coordination.

In this study, we established a feasible strategy to construct a new type of metallo-polymer with helicoidal structure through the combination of covalent polymerization and intramolecular coordination-driven self-assembly. In the design, a tetratopic monomer (M) was prepared with two terminal alkynes in the outer rim for polymerization, and two terpyridines (TPYs) in the inner rim for subsequent folding by selective intramolecular coordination. Then, the linear covalent polymer (P) was synthesized by polymerization of M via Glaser-Hay homocoupling reaction. Finally, intramolecular coordination interactions between TPYs and Zn(II) folded the backbone of P into a right- or left-handed metallo-helicoid (H) with double rims. Owing to multiple positive charges on the inner rim of helicoid, double-stranded DNA molecules (dsDNA) could interact with H through electrostatic interactions. Remarkably, dsDNA allowed exclusive formation of H with right handedness by means of chiral induction.

Overexpression of LINC00261 inhibits non-small cell lung cancer cells progression by interacting with miR-522-3p and suppressing Wnt signaling.

Long noncoding RNA LINC00261 has been experimentally validated to function as a tumor suppressor in several cancers, but its pathological role and functional mechanism in non-small cell lung cancer (NSCLC) are largely unclear. In this study, LINC00261 was delineated in NSCLC to be significantly downregulated in cancer tissues compared with corresponding adjacent normal tissues. Low expression of LINC00261 predicted worse survival for patients with NSCLC. Overexpression of LINC00261 in NSCLC cell lines inhibited cell proliferation and invasion, meanwhile promoted apoptosis. Subcellular fractionation assay showed that LINC00261 existed mainly in the cytoplasm of NSCLC A549 cells and luciferase assay validated its direct interaction with miR-522-3p. Overexpression of miR-522-3p significantly ameliorated suppressive effects of LINC00261 on proliferation and invasion of NSCLC cells. Besides, miR-522-3p was found to be able to directly combine with the 3'-untranslated region of SFRP2, which was generally regarded as a suppressor of Wnt signaling. Further quantitative reverse transcription polymerase chain reaction and Western blot experiments showed that LINC00261 upregulation potentiated the expression of SFRP2 and inhibited Wnt signaling pathway, which could both be reversely modulated by miR-522-3p. Taken together, our study demonstrated that LINC00261 suppressed NSCLC cells progression via sponging miR-522-3p and inhibiting Wnt signaling. These results supported us to better understand the pathogenic mechanism of NSCLC and revealed a potential molecular target for this fatal disease.

Light-Controlled Generation of Singlet Oxygen within a Discrete Dual-Stage Metallacycle for Cancer Therapy.

Noninvasive control over the reversible generation of singlet oxygen (1O2) has found the practical significance in benefiting photodynamic therapy. In this study, we developed a new dual-stage metallacycle (M) by using a photosensitizer and photochromic switch as the functional building blocks, which enables the noninvasive "off-on" switching of 1O2 generation through the efficient intramolecular energy transfer. Due to the proximal placement of the functional entities within the well-defined metallacyclic scaffold, 1O2 generation in the ring-closed form state of the photochromic switch (C-M) is quenched by photoinduced energy transfer, whereas the generation of 1O2 in the ring-open form state (O-M) is activated upon light irradiation. More interestingly, the metallacycle-loaded nanoparticles with relatively high stability and water solubility were prepared, which allow for the delivery of metallacycles to cancer cells via endocytosis. Their theranostic potential has been systematically investigated both in vitro and in vivo. Under the light irradiation, the designed ring-open form nanoparticles (O-NPs) show remarkable higher cytotoxicity against cancer cells compared to the ring-closed form nanoparticles (C-NPs). In vivo experiments also revealed that tumors can be very efficiently eliminated by the designed nanoparticles under light irradiation with the ability to regulate in vivo generation of singlet oxygen. All these results demonstrated that the supramolecular coordination complexes with a dual-stage state provide a highly efficient nanoplatform for noninvasive control over the reversible generation of 1O2, thus allowing for their promising applications in tumor treatment and beyond.

Supramolecular Transformation of Metallacycle-linked Star Polymers Driven by Simple Phosphine Ligand-Exchange Reaction.

As a common phenomenon in biological systems, supramolecular transformations of biomacromolecules lead to specific biological functions as outputs, which thus inspire people to construct biomimetic dynamic systems through supramolecular transformation strategy. It should be noted that well-modulating the artificial macromolecules to fine-tune their properties is of great significance yet still remains a big challenge in polymer chemistry. In this study, through the combination of coordination-driven self-assembly and postassembly ring-opening polymerization, a six-armed star polymer linked by well-defined hexagonal metallacycle as core was successfully prepared. At the same time, the trans-platinum acetylide moieties as transformation sites were anchored onto the discrete metallacycle scaffold. Subsequently, the simple phosphine ligand-exchange reaction induced the conversions of platinum acetylide building blocks with the varied binding angles, which thus resulted in the successive hexagon-rhomboid-hexagon transformations of metallacyclic scaffold, therefore allowing for the corresponding supramolecular transformation of metallacycle-linked star polymers. More importantly, accompanied by such transformation process, property modulation of the resultant polymers has been successfully realized. In a word, by taking advantage of dynamic nature of metal-ligand coordination bonds and simple phosphine ligand-exchange reactions, facile architecture transformation of a star polymer to a linear polymer and back to a star polymer was successfully realized, which may provide a promising approach toward the construction of new dynamic polymeric materials.

Photoactivated Trifunctional Platinum Nanobiotics for Precise Synergism of Multiple Antibacterial Modes.

Integrating multiple strategies of antibacterial mechanisms into one has been proven to have tremendous promise for improving antimicrobial efficiency. Hence, dual-valent platinum nanoparticles (dvPtNPs) with a zero-valent platinum core (Pt0 ) and bi-valent platinum shell (Pt2+ ions), combining photothermal and photodynamic therapy, together with "chemotherapy," emerge as spatiotemporally light-activatable platinum nano-antibiotics. Under near-infrared (NIR) exposure, the multiple antibacterial modes of dvPtNPs are triggered. The Pt0 core reveals significant hyperthermia via effective photothermal conversion while an immediate release of chemotherapeutic Pt2+ ions occurs through hyperthermia-initiated destabilization of metallic interactions, together with reactive oxygen species (ROS) level increase, thereby resulting in synergistic antibacterial effects. The precise cooperative effects between photothermal, photodynamic, and Pt2+ antibacterial effects are achieved on both Gram-negative Escherichia coli and Gram-positive methicillin-resistant Staphylococcus aureus, where bacterial viability and colony-forming units are significantly reduced. Moreover, similar results are observed in mice subcutaneous abscess models. Significantly, after NIR treatment, dvPtNP exhibits a more robust bacteria-killing efficiency than other PtNP groups, owing to its integration of dramatic damage to the bacterial membrane and DNA, and alteration to ATP and ROS metabolism. This study broadens the avenues for designing and synthesizing antibacterial materials with higher efficiency.

Photoresponsive Chirality-Tunable Supramolecular Metallacycles.

Chirality-tunable supramolecular metallacycles containing two light-responsive dithienylethene units and two chiral 1,1'-bi-2-naphthol (BINOL) units have been successfully constructed via coordination-driven self-assembly. These new metallacycles are well-characterized with 1D multinuclear NMR (1 H and 31 P NMR), 2D 1 H-1 H COSY and DOSY, ESI-TOF-MS, and PM6 semiempirical molecular orbital methods. Interestingly, upon irradiation with ultraviolet and visible light, the conformation of these metallacycles can undergo reversible transformation between ring-open and ring-closed forms accompanied with the obvious change of CD signals. Further investigation reveals that the photoisomerization of the dithienylethene moieties induces the change in the dihedral angle of the binaphthyl rings, thus leading to the chiral modulation of supramolecular metallacycles. Thus, this study provides very few examples of the light-induced chirality-tunable metallosupramolecular assemblies, which may find potential application in mimicking the function of natural systems in the future.

CO2 Stimuli-Responsive, Injectable Block Copolymer Hydrogels Cross-Linked by Discrete Organoplatinum(II) Metallacycles via Stepwise Post-Assembly Polymerization.

Supramolecular polymeric gels cross-linked by well-defined, discrete metal-organic macrocycles (MOMs) or metal-organic cages have become a prevailing topic within the field of supramolecular self-assembly. However, the realization of supramolecular polymeric hydrogels cross-linked by discrete organometallic architectures with good biocompatibility is still a great challenge. Herein, we present the successful preparation of CO2 stimuli-responsive, injectable block copolymer hydrogels cross-linked by discrete organoplatinum(II) metallacycles. Through the combination of coordination-driven self-assembly and stepwise post-assembly polymerization, star block copolymers (SBCPs) containing well-defined hexagonal metallacycles as cores were successfully prepared, which featured CO2 stimuli-responsive properties including CO2-triggered morphology transition and CO2-induced thermoresponsive behavior. Interestingly, the resultant SBCPs were capable of forming supramolecular hydrogels with MOMs as junctions near physiological temperature, which allowed the realization of a reversible gel-to-sol transformation through the removal and addition of CO2. More importantly, the resultant supramolecular hydrogels presented good cytocompatibility in vitro. Therefore, this study provides a new strategy for the construction of new "smart" supramolecular hydrogels with promising applications as biological materials.

Optimizing Sugarcane Clonal Propagation In Vitro by Using Calcium Ammonium Nitrate and Ammonium Sulfate.

To replace explosive nitrate-based chemicals in MS medium, this study developed a new, safer, and more cost-effective method using fertilizer-grade calcium ammonium nitrate and ammonium sulfate. This approach replaces ammonium nitrate and potassium nitrate, ensuring both safety and cost efficiency for sugarcane propagation. Six local sugarcane varieties-Zhongtang1 (ZT1), Zhongtang3 (ZT3), Zhongtang6 (ZT6), Guitang42 (GT42), Guitang44 (GT44), and Guiliu 07150 (GT07150)-were used. In the control group (Ck), nitrate ions (NO3-) were 39.28 mM, and ammonium ions (NH4+) were 20.49 mM, with a 2:1 ratio. In the treatment groups, the concentrations of nitrate ions (NO3-) and ammonium ions (NH4+) included treatment 1 (19.69 mM NO3- and 10.3 mM NH4+), treatment 2 (29.54 mM and 15.44 mM), treatment 3 (39.38 mM and 20.59 mM), treatment 4 (49.225 mM and 25.74 mM), treatment 5 (59.07 mM and 30.89 mM), and treatment 6 (68.915 mM and 36.03 mM), respectively, all with the same 2:1 ratio. Fifty bottles per treatment, with three replicates, were used for each sugarcane plantlets treatment. After five subcultures, the optimal ratio was determined by assessing morphological and physiological parameters, nitrogen levels, and SOD enzyme activity. The results indicated that treatment 3 (39.38 mM and 20.59 mM) and treatment 4 (49.225 mM and 25.74 mM) had the best morphological and physiological indicators. The optimal doses of calcium ammonium nitrate and ammonium sulfate were found in treatments 3 and 4, as well as in the control, with no significant difference among them. However, treatment 3, due to its lower dose, was more cost effective. To improve cost efficiency in practical production, it is recommended to use the lower concentration ratio of treatment 3 for plant tissue culture plantlets.

A Novel Homozygous Missense Mutation (Ile583Asn) in a Consanguineous Marriage Family with Hereditary Factor XII Deficiency: A Case Report.

BACKGROUND: Hereditary coagulation factor XII (FXII) deficiency is an autosomal recessive disorder. At present, the contribution of severe FXII deficiency to the development of thromboembolism is still undetermined. There are limited reports on the relationship between the FXII defect and thromboembolism. CASE PRESENTATION: A 27-year-old woman came to our hospital for the treatment of shoulder trauma and cervical disc herniation caused by a car accident. The shoulder trauma was treated with five stitches. After physical examination, imaging examination, and routine coagulation examination, cervical disc herniation was treated conservatively. Combined with the examination results, the patient was diagnosed with FXII deficiency. Unfortunately, the patient was readmitted 10 days after the trauma with edema in the lower limbs and secondary varicose veins. The D-dimer increased to 6.22 mg/L. Thrombus in the inferior vena cava and right common iliac was shown by lower limb venography. According to the patient's medical history, the F12 gene was analyzed by direct sequencing. The patient was also screened for other thrombotic risk factors. Genetic analysis showed that the patient had a c.1748T > A (p.Ile583Asn) homozygous missense mutation in exon 14 of the F12 gene. No other hereditary thrombophilia risk factors screened were positive in the patient. CONCLUSION: The p.Ile583Asn missense mutation in exon 14 of the F12 gene might be responsible for the reduction of the FXII level in the patient.

Synthesis and Characterization of a Novel Composite Edible Film Based on Hydroxypropyl Methyl Cellulose Grafted with Gelatin.

A novel composite edible film was synthesized by grafting gelatin chain onto hydroxypropyl methyl cellulose (HPMC) in the presence of glycerol (used as a plasticizer) using a solution polymerization technique. The reaction was carried out in homogeneous aqueous medium. Thermal properties, chemical structure, crystallinity, surface morphology, and mechanical and hydrophilic performance changes of HPMC caused by the addition of gelatin were investigated by differential scanning calorimetry, thermogravimetric, Fourier transform infrared spectroscopy, scanning electron microscopy, X-ray diffraction, universal testing machine and water contact angle. The results shows that HPMC and gelatin are miscible and the hydrophobic property of the blending film can be enhanced with the introduction of the gelatin. Moreover, the HPMC/gelatin blend films are flexible, and exhibit excellent compatibility, good mechanical properties and also thermal stability, and could be promising candidates for food packaging materials.