Dynamic Control of Cyclic Peptide Assembly to Form Higher-Order Assemblies.

Chirality correction, asymmetry, ring-chain tautomerism and hierarchical assemblies are fundamental phenomena in nature. They are geometrically related and may impact the biological roles of a protein or other supermolecules. It is challenging to study those behaviors within an artificial system due to the complexity of displaying these features. Herein, we design an alternating D,L peptide to recreate and validate the naturally occurring chirality inversion prior to cyclization in water. The resulting asymmetrical cyclic peptide containing a 4-imidazolidinone ring provides an excellent platform to study the ring-chain tautomerism, thermostability and dynamic assembly of the nanostructures. Different from traditional cyclic D,L peptides, the formation of 4-imidazolidinone promotes the formation of intertwined nanostructures. Analysis of the nanostructures confirmed the left-handedness, representing chirality induced self-assembly. This proves that a rationally designed peptide can mimic multiple natural phenomena and could promote the development of functional biomaterials, catalysts, antibiotics, and supermolecules.

Pyrroloquinoline quinone ameliorates liver injury in mice induced by cyclophosphamide.

The current study aimed to investigate the potential ameliorative effects of pyrroloquinoline quinone (PQQ) on cyclophosphamide (CTX)-induced liver injury in mice. The liver injury model was established by injecting mice with CTX (80 mg/kg/day). Liver function indices, antioxidant enzyme activities, and inflammatory cytokines were evaluated. In addition, protein expression levels of the nuclear factor E2-related factor 2 (Nrf2) and nuclear factor kappa-B (NF-κB) pathways in the liver tissues were determined using western blot. The results indicated that PQQ decreased the serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and the malondialdehyde (MDA), interleukin (IL)-1ß, IL-6, tumor necrosis factor-α (TNF-α) levels in the liver tissues. Moreover, PQQ enhanced the activities of oxidative stress markers to alleviate CTX induced oxidative stress. Furthermore, the expression levels of heme oxygenase-1 (HO-1), glutamate-cysteine ligase modifier subunit (GCLM), and NAD(P)H quinone oxidoreductase 1 (NQO1) were significantly increased, and the expression levels of NF-κB p50, NF-κB p65, and inhibitor of NF-κB kinase alpha (IKKα) were significantly decreased after PQQ administration, suggesting that PQQ alleviated CTX-induced liver injury via activating the Nrf2-mediated antioxidant response pathway, and inhibiting the NF-κB-mediated inflammation pathway. Therefore, PQQ can be potentially used as a dietary supplement or functional foods for alleviating the CTX-induced liver injury.

Structure-Based Programming of Supramolecular Assemblies in Living Cells for Selective Cancer Cell Inhibition.

Here we report on the design, synthesis, and assembly of an enzymatic programmable peptide system inspired by endocytic processes to induce molecular assemblies formation spatiotemporally in living cancer cells, resulting in glioblastoma cell death mainly in necroptosis. Our results indicate the stability and glycosylation of molecules play an essential role in determining the final bioactivity. Detailed mechanistic studies by CLSM, Flow cytometry, western blot, and Bio-EM suggest the site-specific formation of assemblies, which could induce the LMP and activate the downstream cell death pathway. Moreover, we also demonstrate that our strategy can boost the activity of commercial chemotherapy drug by escaping lysosome sequestration. We expected this work would be expanded towards artificial intelligent biomaterials for cancer therapy and imaging precisely.

Improved Stabilities of Labeling Probes for the Selective Modification of Endogenous Proteins in Living Cells and In Vivo.

To date, various affinity-based protein labeling probes have been developed and applied in biological research to modify endogenous proteins in cell lysates and on the cell surface. However, the reactive groups on the labeling probes are also the cause of probe instability and nonselective labeling in a more complex environment, e. g., intracellular and in vivo. Here, we show that labeling probes composed of a sterically stabilized difluorophenyl pivalate can achieve efficient and selective labeling of endogenous proteins on the cell surface, inside living cells and in vivo. As compared with the existing protein labeling probes, probes with the difluorophenyl pivalate exhibit several advantages, including long-term stability in stock solutions, resistance to enzymatic hydrolysis and can be customized easily with diverse fluorophores and protein ligands. With this probe design, endogenous hypoxia biomarker in living cells and nude mice were successfully labeled and validated by in vivo, ex vivo, and immunohistochemistry imaging.

Self-Immolative Difluorophenyl Ester Linker for Affinity-Based Fluorescence Turn-on Protein Detection.

Currently most fluorogenic probes are developed for the analysis of enzymes, where a bond breaking or rearrangement reaction is required to transform a nonfluorescent enzymatic substrate into a fluorescent product. However, this approach cannot be used for proteins that do not possess enzymatic activities. In this article, we show that fluorogenic probes with a self-immolative difluorophenyl ester linker can mimic the bond disassembly processes of fluorogenic enzyme substrates for the rapid analysis of nonenzymatic proteins. Although numerous self-immolative reagents have shown promising applications in sensors, drug delivery systems, and material chemistry, all of them are triggered by either enzymes or small reactive molecules. In our strategy, the probe binds to the protein via a specific protein-ligand interaction, inducing a chemical reaction between the self-immolative linker and an amino acid of the protein, thereby triggering a cascade reaction that leads to the activation and release of the fluorogenic reporter. In contrast, a phenyl ester linker without the difluoro substituent cannot be triggered to release the fluorogenic reporter. With this probe design, live-cell imaging of extracellular and intracellular endogenous tumor marker proteins can be achieved with high selectivity and sensitivity.

Rapid and Selective Labeling of Endogenous Transmembrane Proteins in Living Cells with a Difluorophenyl Ester Affinity-Based Probe.

The long-term stability of affinity-based protein labeling probes is crucial to obtain reproducible protein labeling results. However, highly stable probes generally suffer from low protein labeling efficiency and pose significant challenges when labeling low abundance native proteins in living cells. In this paper, we report that protein labeling probes based on an ortho-difluorophenyl ester reactive module exhibit long-term stability in DMSO stock solution and aqueous buffer, yet they can undergo rapid and selective labeling of native proteins. This novel electrophile can be customized with a wide range of different protein ligands and is particularly well-suited for the labeling and imaging of transmembrane proteins. With this probe design, the identity and relative levels of basal and hypoxia-induced transmembrane carbonic anhydrases were revealed by live cell imaging and in-gel fluorescence analysis. We believe that the extension of this difluorophenyl ester reactive module would allow for the specific labeling of various endogenous membrane proteins, facilitating in-depth studies of their distribution and functions in biological processes.

Ameliorate effect of pyrroloquinoline quinone against cyclophosphamide-induced nephrotoxicity by activating the Nrf2 pathway and inhibiting the NLRP3 pathway.

AIMS: Cyclophosphamide (CTX) is an effective anti-tumor and immunosuppressive agent, but it induces nephrotoxicity in clinical applications. The present study aimed to evaluate the protective effect of pyrroloquinoline quinone (PQQ) on CTX-induced nephrotoxicity. MAIN METHODS: We injected male ICR mice with CTX (80 mg/kg/day), and determined nephrotoxicity indices, MDA and antioxidant defenses, inflammatory cytokines, and the levels of main proteins in the Nrf2-HO-1 and NLRP3 signaling pathways. KEY FINDINGS: PQQ has significantly decreased the serum levels of creatinine and urea compared to Model group. When treated with PQQ, MDA, IL-1ß, IL-6, and TNF-α levels have decreased, and SOD, GSH-Px, and CAT activity have increased in the kidney tissues of CTX-induced mice. PQQ activated the Nrf2-mediated signaling pathway, as indicated by the increased expression of Nrf2, HO-1, GCLM, and NQO1. Moreover, PQQ inhibited the NLRP3 inflammatory pathway, as indicated by the reduced expression of NLRP3, ASC, and Caspase-1. SIGNIFICANCE: Our results suggest that PQQ protects against CTX-induced nephrotoxicity, probably by activating the Nrf2-mediated antioxidant pathway and inhibiting the NLRP3 inflammatory pathway.

Collagen Extracted from Bigeye Tuna (Thunnus obesus) Skin by Isoelectric Precipitation: Physicochemical Properties, Proliferation, and Migration Activities.

Collagen was extracted from bigeye tuna (Thunnus obesus) skins by salting-out (PSC-SO) and isoelectric precipitation (PSC-IP) methods. The yield of the PSC-IP product was approximately 17.17% (dry weight), which was greater than the yield obtained from PSC-SO (14.14% dry weight). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis indicated that collagen from bigeye tuna skin belongs to collagen type I. Inductively coupled plasma mass spectrometry results indicate that the heavy metal abundance in PSC-IP was lower than the maximum acceptable amounts according to Chinese regulatory standards. In addition, results from a methylthiazolyldiphenyl-tetrazolium bromide assay and an in vitro scratch assay demonstrated that PSC-IP could promote the proliferation and migration of NIH-3T3 fibroblasts. Overall, results suggest PSC-IP could be used to rapidly extract collagen from marine by-products instead of traditional salting-out methods. Collagen from bigeye tuna skin may also have strong potential for cosmetic and biomedical applications.

Novel method to measure enamel surface porosity with hydrogen peroxide bleaching.

PURPOSE: To establish methods to quantitate the physical surface change, not the chemical color bleaching change, of enamel occurring with hydrogen peroxide treatment in solution and commercially available plastic strips. METHODS: Bovine enamel was polished to a sound, uniform, optical flat, white subsurface that was used as the initial substrate for all substrate modification, treatment and instrumental measurement using digital photography-image analysis, SEM and profilometry. Sound enamel was treated with 10, 20 and 30% solutions of hydrogen peroxide. Plastic strips were used to treat both sound and acid modified enamel surfaces. Etched enamel, similar to a 10-second exposure to lemon juice, was treated with 0.5, 1, 3, 5, 10, and 15% hydrogen peroxide for 24-hour exposure at 37 degrees C to obtain a dose response curve to this modified enamel. RESULTS: The digital photography-image analysis system and scanning electron microscopy (SEM) were effective in detecting enamel surface porosity and structure disruptive changes, respectively. Plastic strip treatment of both sound and etched enamel produced little surface change. Measurable surface change of etched enamel was detected with as low as 1% hydrogen peroxide in solution. The surface change with 15% hydrogen peroxide was statistically significant. Dye uptake as measured by image analysis indicated an increase in surface porosity that was more evident with the acid modified surface. SEM studies were consistent with this observation.

[Effects of hyperthermia on brainstem auditory evoked potentials and middle latency response in rats].

AIM: To study the effects of hyperthermia on brainstem auditory evoked potentials (BAEP) and middle latency response (MLR) in rats. METHODS: BAEP and MLR were recorded at the skull surface of rats. The body temperature of anesthetized rats increased gradually with a physical method and was detected by a digital thermometer inserted into the rectum. The peak latency (PL), interpeak latency (IPL), wave amplitude and the critical body temperature at which BAEP and MLR completely lost had been observed. RESULTS: All PL and I - II, I - III and I -IV IPL of BAEP shortened more and more as the body temperature increased step by step from 37 degrees C to 41.5 degrees C. But all PL and I - II and I -IV IPL did not shortened further and prolonged a little contrary as the body temperature at 42 degrees C and over 42 degrees C. All PL and P1-P3 and P2-P3 IPL of MLR also shortened as the body temperature increased from 37 degrees C to 43 degrees C. The wave amplitudes of BAEP and MLR decreased as the body temperature increased, especially as the body temperature over 42 degrees C. BAEP and MLR lost completely and synchronously at the body temperature (43.1 +/- 0.5) degrees C, which was not reversed as the body temperature returning to normal by cooling. CONCLUSION: There were obvious effects of hyperthermia on both BAEP and MLR in rats, and irreversible impairments appeared at a critical body temperature.