Sedimentary organic molecular compositions reveal the influence of glacier retreat on ecology on the Tibetan Plateau.

Global warming and glacier retreat have significant impacts on the structure and function of natural ecosystems. However, little is known about how glacier retreat affects the long-term evolution of ecosystems at high-altitude regions. In this study, we explored the possible effects of glacier retreat on catchment vegetation and lake productivity in Lake Puma Yumco, southeastern Tibetan Plateau, based on detailed organic molecular compositions determined by an ultra-high-resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS), and combined with various sedimentary geochemical indicators. The glaciers in the catchment keep retreating since 1870 CE, as inferred from the multiple indices of total organic carbon content (TOC), total nitrogen content (TN), C/N ratios, and carbonate contents. Accompanying modern global warming and glacier shrinkage, the relative abundance of soil- and vegetation-derived large molecular compounds (e.g., vascular plant-derived polyphenols, highly unsaturated and phenolic compounds, and condensed aromatics) increased gradually in lake sediments, suggesting that ice-covered land was exposed under warming condition, and gradually revegetation occurred. Both increases in relative abundance of nitrogen-containing compounds (e.g., CHNO) and chlorophyll derivative contents in the lake sediments were observed since 1870 CE, suggesting that stronger catchment weathering and increasing terrestrial nutrient loads enhanced the downstream lake productivity after glacier retreat. Our results imply that continued global warming and alpine glacier retreat in the future may further promote vegetation expansion and increases in lake productivity on the Tibetan Plateau.

Sedimentary organic matter molecular composition reveals the eutrophication of the past 500 years in Lake Daihai, Inner Mongolia.

Lake eutrophication seriously threatens water quality and human health. Under continuous global warming and intensified human activity, increasing attention is being paid to how lake trophic status responds to climate change and anthropogenic impacts. Based on the sedimentary organic matter (SOM) molecular composition determined by the Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) technology, and combined with the SOM stable nitrogen isotopes (δ15Norg), we studied how lake trophic status and ecology respond to both climatic changes and anthropogenic impacts of the past 500 yrs at Lake Daihai, Inner Mongolia. The results show that the relative abundance of lipids, proteins, and carbohydrates in lake sediments kept relatively low before AD ∼1850, and increased gradually thereafter, especially after AD ∼1950, suggesting that the lake trophic status was low before AD ∼1850, but obviously increased during the past one more century. On the other hand, the relative abundance of allochthonous condensed aromatics and vascular plant-derived polyphenols compounds gradually decreased after AD ∼1850, which is most likely due to the intensified land-use changes in the catchment. Our results show that the SOM molecular composition is more sensitive to trace the land-use changes than the δ15Norg ratios, suggesting a potential use of this technique to trace even earlier human land uses (e.g., during the prehistorical times) in a catchment. The results of this study suggest that intensified land-use change, increased discharges of human sewage and industrial wastewater, cropland runoff, and concentrated effects caused by lake level drops may have combinedly increased nutrient concentration and accelerated lake eutrophication at Lake Daihai. Therefore, proper policy is necessary to slow down anthropogenic impacts and limit further eutrophication for lakes like Lake Daihai.

DNAzyme recognition triggered cascade signal amplification for rapid and highly sensitive visual detection of uranyl ions.

This work presents a rapid and highly sensitive colorimetric assay using bifunctional DNA probe decorated agarose microbeads (MBs) coupled with a cascade signal amplification system, including rolling circle amplification (RCA) and the hemin/G-quadruplex-catalyzed colorimetric reaction, for visualized detection of uranyl ions. The DNA probe integrates the UO22+-specific DNAzyme/substrate as the target recognition unit and a DNA primer as the signal conversion unit. The presence of uranyl ions induces the efficient cleavage of the DNA substrates with the catalysis of DNAzyme. Then the conjugated primers are released from MBs, initiating the RCA reaction (the first amplification). The RCA product consists of repetitive G-quadruplexes that can lead to a second amplification by catalyzing the oxidation of ABTS2- with hemin binding, resulting in a coloration that is visible to the naked eye. The whole assay procedure could be finished within 40 min, including recognition of uranyl and DNA cleavage (5 min), the RCA reaction (30 min) and data readout either by eye or using a UV-vis spectrometer (5 min for each sample). In the optimal conditions, concentrations as low as 5 nM uranyl ions could be distinguished by the naked eye. With UV-vis spectrometric measurement, the visible absorbance had a linear relationship with the concentration of uranyl ions with a dynamic range from 1 nM to 50 nM, and a low detection limit of 0.48 nM (i.e. ∼0.12 ppb) was obtained. Excellent selectivity and anti-interference capability in water samples were also certified. This facile visualized assay could be applied in detecting trace-level uranium for on-site environmental analysis.

A general controllable release amplification strategy of liposomes for single-particle collision electrochemical biosensing.

As an important component of the COVID-19 mRNA vaccines, liposomes play a key role in the efficient protection and delivery of mRNA to cells. Herein, due to the controllable release amplification strategy of liposomes, a reliable and robust single-particle collision electrochemical (SPCE) biosensor was constructed for H9N2 avian influenza virus (H9N2 AIV) detection by combining liposome encapsulation-release strategy with immunomagnetic separation. The liposomes modified with biotin and loaded with platinum nanoparticles (Pt NPs) were used as signal probes for the first time. Biotin facilitated the coupling of biomolecules (DNA or antibodies) through the specific reaction of biotin-streptavidin. Each liposome can encapsulate multiple Pt NPs, which were ruptured under the presence of 1 × PBST (phosphate buffer saline with 0.05% Tween-20) within 2 min, and the encapsulated Pt NPs were released for SPCE experiment. The combination of immunomagnetic separation not only improved the anti-interference capabilities but also avoided the agglomeration of Pt NPs, enabling the SPCE biosensor to realize ultrasensitive detection of 18.1 fg/mL H9N2 AIV. Furthermore, the reliable SPCE biosensor was successfully applied in specific detection of H9N2 AIV in complex samples (chicken serum, chicken liver and chicken lung), which promoted the universality of SPCE biosensor and its application prospect in early diagnosis of diseases.

Cloud point extraction associated with differential pulse voltammetry: preconcentration and determination of trace uranyl in natural water.

A procedure for the electroanalytical determination of uranyl ions pre-concentrated from natural water by cloud point extraction (CPE) is developed in this study. CPE parameters, such as surfactant concentration, extractant concentration, pH and additive concentration were optimized. After CPE, the solution was diluted for electrochemical determination by differential pulse voltammetry (DPV) with a mercury film electrode (Hg-GCE). The current response of uranyl showed a linear relationship with concentration from 10 nmol L-1 to 1 µmol L-1. The hyphenated method combining CPE and DPV achieved a detection limit of uranyl as low as 0.15 nmol L-1. The presence of some foreign ions interfered greatly with the current response of electrochemical detection. Therefore, the hyphenated technique combining CPE and DPV is important because the CPE step provides selectivity against the co-existing metal ions for electrochemical detection. No interference was seen from the representative foreign metal ions in the CPE-DPV method. The developed method was successfully applied for the determination of uranyl ions in natural water. The average recovery using CPE-DPV in real samples varied from 94.4% to 103.2% and the precision was comparable with that of inductively coupled plasma mass spectrometry (ICP-MS), indicating the good accuracy and precision of the method developed. This hyphenated technique could have greater potential applications for the determination of uranyl ions in aqueous environments.

Co-assembled nanocomplexes of peptide neoantigen Adpgk and Toll-like receptor 9 agonist CpG ODN for efficient colorectal cancer immunotherapy.

Cancer vaccines targeting tumor specific neoantigens derived from nonsynonymous mutations of tumor cells have emerged as an effective approach to induce antitumor T cells responses for personalized cancer immunotherapy. Despite the enormous potential of synthetic peptides as a common modality for neoantigen vaccines, their practical efficacy was limited due to their relatively low immunogenicity. Herein, we modify neoantigen peptide (Adpgk) derived from MC-38 colon carcinoma by supplementing ten consecutive positively-charged lysines (10 K-Adpgk) to obtain cationic polypeptide. And then we made them self-assemble with toll-like receptor 9 (TLR-9) agonist CpG oligodeoxynucleotides (CpG ODN) adjuvant directly forming antigen/adjuvant integrated nanocomplexes (PCNPs) through electrostatic interaction for potent tumor immunotherapy. The optimal formed PCNPs were around 175 nm with uniform size distribution and could maintain stability in physiological saline solution. CpG ODN and 10 K-Adpgk in the formed PCNPs could be effectively uptake by dendritic cells (DCs) and stimulate the maturation of DCs as well as improving the efficiency of antigen cross-presentation efficiency in vitro. Furthermore, the PCNPs vaccine could markedly improve neoantigen and adjuvant co-delivery efficiency to lymphoid organs and activate cytotoxic T cells. In addition, vaccination with PCNPs could not only offer prophylactic to protect mice from challenged MC-38 colorectal tumors, but also achieve a better anti-tumor effect in an established colorectal tumor model, and significantly prolong the survival rate of tumor-bearing mice. Therefore, this work provided a versatile but effective method for neoantigen peptide and CpG ODN co-assembly vaccine platform for efficient colorectal cancer immunotherapy.

In Situ Surface-Enhanced Raman Spectroscopy Detection of Uranyl Ions with Silver Nanorod-Decorated Tape.

Surface-enhanced Raman spectroscopy (SERS) has been utilized for rapid analysis of uranyl ions (UO2 2+) on account of its fast response and high sensitivity. However, the difficulty of fabricating a suitable SERS substrate for in situ analysis of uranyl ions severely restricts its practical application. Hence, we proposed flexible and adhesive SERS tape decorated with silver nanorod (AgNR) arrays for in situ detection of UO2 2+. The SERS tape was fabricated through a simple "paste & peel off" procedure by transferring the slanted AgNR arrays from silicon to the transparent tape surface. UO2 2+ can be easily in situ detected by placing the AgNR SERS tape into an aqueous solution or pasting it onto the solid matrix surface due to the excellent transparent feature of the tape. The proposed SERS tape with well-distributed AgNRs effectively improved the reproducibility and sensitivity for UO2 2+ analysis. UO2 2+ with concentration as low as 100 nM was easily detected. Besides, UO2 2+ adsorbed on an iron disc and rock surface also can be rapidly in situ detected. With its simplicity and convenience, the AgNR SERS tape-based SERS technique offers a promising approach for environmental monitoring and nuclear accident emergency detection.

Purification of inclusion bodies using PEG precipitation under denaturing conditions to produce recombinant therapeutic proteins from Escherichia coli.

It has been documented that the purification of inclusion bodies from Escherichia coli by size exclusion chromatography (SEC) may benefit subsequent refolding and recovery of recombinant proteins. However, loading volume and the high cost of the column limits its application in large-scale manufacturing of biopharmaceutical proteins. We report a novel process using polyethylene glycol (PEG) precipitation under denaturing conditions to replace SEC for rapid purification of inclusion bodies containing recombinant therapeutic proteins. Using recombinant human interleukin 15 (rhIL-15) as an example, inclusion bodies of rhIL-15 were solubilized in 7 M guanidine hydrochloride, and rhIL-15 was precipitated by the addition of PEG 6000. A final concentration of 5% (w/v) PEG 6000 was found to be optimal to precipitate target proteins and enhance recovery and purity. Compared to the previously reported S-200 size exclusion purification method, PEG precipitation was easier to scale up and achieved the same protein yields and quality of the product. PEG precipitation also reduced manufacturing time by about 50 and 95% of material costs. After refolding and further purification, the rhIL-15 product was highly pure and demonstrated a comparable bioactivity with a rhIL-15 reference standard. Our studies demonstrated that PEG precipitation of inclusion bodies under denaturing conditions holds significant potential as a manufacturing process for biopharmaceuticals from E. coli protein expression systems.

A novel GLP-1 analog, a dimer of GLP-1 via covalent linkage by a lysine, prolongs the action of GLP-1 in the treatment of type 2 diabetes.

GLP-1 is an incretin hormone that can effectively lower blood glucose, however, the short time of biological activity and the side effect limit its therapeutic application. Many methods have been tried to optimize GLP-1 to extend its in vivo half-time, reduce its side effect and enhance its activity. Here we have chosen the idea to dimerize GLP-1 with a C-terminal lysine to form a new GLP-1 analog, DLG3312. We have explored the structure and the biological property of DLG3312, and the results indicated that DLG3312 not only remained the ability to activate the GLP-1R, but also strongly stimulated Min6 cell to secrete insulin. The in vivo bioactivities have been tested on two kinds of animal models, the STZ induced T2DM mice and the db/db mice, respectively. DLG3312 showed potent anti-diabetic ability in glucose tolerance assay and single-dose administration of DLG3312 could lower blood glucose for at least 10 hours. Long-term treatment with DLG3312 can reduce fasted blood glucose, decrease water consumption and food intake and significantly reduce the HbA1c level by 1.80% and 2.37% on STZ induced T2DM mice and the db/db mice, respectively. We also compared DLG3312 with liraglutide to investigate its integrated control of the type 2 diabetes. The results indicated that DLG3312 almost has the same effect as liraglutide but with a much simpler preparation process. In conclusion, we, by using C-terminal lysine as a linker, have synthesized a novel GLP-1 analog, DLG3312. With simplified preparation and improved physiological characterizations, DLG3312 could be considered as a promising candidate for the type 2 diabetes therapy.

A novel self-cleavable tag Zbasic-∆I-CM and its application in the soluble expression of recombinant human interleukin-15 in Escherichia coli.

Soluble expression of recombinant therapeutic proteins in Escherichia coli (E. coli) has been a challenging task in biopharmaceutical development. In this study, a novel self-cleavable tag Zbasic-intein has been constructed for the soluble expression and purification of a recombinant cytokine, human interleukin-15 (IL-15). We screened several solubilizing tags fused with the self-cleavable Mycobacterium tuberculosis recA mini-intein ∆I-CM and demonstrated that Zbasic tag can significantly improve the solubility of the product with correspondent to the intein activity. The fusion protein "Zbasic-∆I-CM-IL-15" was expressed with high solubility and easily enriched by the cost-effective cation-exchange chromatography. The self-cleavage of the fusion tag Zbasic-∆I-CM was then induced by a pH shift, with an activation energy of 7.48 kcal/mol. The mature IL-15 with natural N-terminus was released and further purified by hydrophobic interaction and anion-exchange chromatography. High-resolution reverse-phase high-performance liquid chromatography and mass spectrometry analysis confirmed that the product was of high purity and correct mass. With a CTLL-2 cell proliferation-based assay, the EC50 was evaluated to be of about 0.126 ng/mL, similar to the product in clinical trials. By avoiding the time-consuming denaturing-refolding steps in previously reported processes, the current method is efficient and cost-effective. The novel tag Zbasic-∆I-CM can be potentially applied to large-scale manufacturing of recombinant human cytokines as well as other mammalian-sourced proteins in E. coli.

The coordination of amidoxime ligands with uranyl in the gas phase: a mass spectrometry and DFT study.

Sequestering uranium from the ocean is a promising solution to fulfill the demand for nuclear energy. Motivated by this purpose, a series of amidoxime ligands and their analogs have been developed with high absorption capacity and selectivity. An in-depth understanding of the structural information of the uranyl-ligand complexes is essential to improve the performance of the ligands. Herein, we have studied the coordination of three amidoxime ligands (6-methoxyl-naphtha-2-amidoxime, NAO; glutarimidedioxime, GIO; and gluardiamidoxime, GDO) with uranyl in the gas phase by mass spectrometry. The identifications of the electrospray ionization (ESI) generated species, the fragmentation pathways upon dissociation, the relative binding affinities of the ligands, and the hydration reactions have been conducted and compared to reveal their structural information in the gas phase. The binding modes for all the complexes were suggested based on the experimental results and were further studied by density functional theory (DFT) calculations.