Freeze substitution Hi-C,a convenient and cost-effective method for capturing the natural 3D chromatin conformation from frozen samples

Chromatin interactions functionally affect genome architecture and gene regulation,but to date,only fresh samples must be used in High-through chromosome conformation capture(Hi-C)to keep natural chromatin conformation intact.This requirement has impeded the advancement of 3 D genome research by limiting sample collection and storage options for researchers and severely limiting the number of samples that can be processed in a short time.Here,we develop a freeze substitution Hi-C(FS-Hi-C)technique that overcomes the need for fresh samples.FS-Hi-C can be used with samples stored in liquid nitrogen(LN2):the water in a vitreous form in the sample cells is replaced with ethanol via automated freeze substitution.After confirming that the FS step preserves the natural chromosome conformation during sample thawing,we tested the performance of FS-Hi-C with Drosophila melanogaster and Gossypium hirsutum.Beyond allowing the use of frozen samples and confirming that FS-Hi-C delivers robust data for generating contact heat maps and delineating A/B compartments and topologically associating domains,we found that FS-HiC outperforms the in situ Hi-C in terms of library quality,reproducibility,and valid interactions.Thus,FS-HiC will probably extend the application of 3D genome structure analysis to the vast number of experimental contexts in biological and medical research for which Hi-C methods have been unfeasible to date.

Drug-peptide supramolecular hydrogel boosting transcorneal permeability and pharmacological activity via ligand-receptor interaction

Boosting transcorneal permeability and pharmacological activity of drug poses a great challenge in the field of ocular drug delivery.In the present study,we propose a drug-peptide supramolecular hydrogel based on anti-inflammatory drug,dexamethasone(Dex),and Arg-Gly-Asp(RGD)motif for boosting transcorneal permeability and pharmacological activity via the ligand-receptor interaction.The drug-peptide(Dex-SA-RGD/RGE)supramolecular hydrogel comprised of uniform nanotube architecture formed spontaneously in phosphate buffered saline(PBS,pH=7.4)without external stimuli.Upon storage at 4℃,25℃,and 37℃ for 70 days,Dex-SA-RGD in hydrogel did not undergo significant hydrolysis,suggesting great long-term stability.In comparison to Dex-SA-RGE,Dex-SA-RGD exhibited a more potent in vitro anti-inflammatory efficacy in lipopolysaccharide(LPS)-activated RAW 264.7 macrophages via the inhibition of nuclear factorкB(NF-κB)signal pathway.More importantly,using drug-peptide supramolecular hydrogel labeled with 7-nitro-2,1,3-benzoxadiazole(NBD),the Dex-SA-K(NBD)RGD showed increased performance in terms of integrin targeting and cellular uptake compared to Dex-SA-K(NBD)RGE,as revealed by cellular uptake assay.On topical instillation in rabbit’s eye,the proposed Dex-SA-K(NBD)RGD could effectively enhance the transcorneal distribution and permeability with respect to the Dex-SA-K(NBD)RGE.Overall,our findings demonstrate the performance of the ligand-receptor interaction for boosting transcorneal permeability and pharmacological activity of drug.