Table_1_Transition patterns of metabolism-weight phenotypes over time: A longitudinal study using the multistate Markov model in China.DOCX

BackgroundA change in weight or metabolic status is a dynamic process, yet most studies have focused on metabolically healthy obesity (MHO) and the transition between MHO and metabolically unhealthy obesity (MUO); therefore, they have not fully revealed the nature of all possible transitions among metabolism-weight phenotypes over the years.MethodsThis was a longitudinal study based on a retrospective health check-up cohort. A total of 9,742 apparently healthy individuals aged 20–60 years at study entry were included and underwent at least two health check-ups. Six metabolism-weight phenotypes were cross-defined by body mass index (BMI) categories and metabolic status as follows: metabolically healthy normal weight (MHNW), metabolically healthy overweight (MHOW), MHO, metabolically unhealthy normal weight (MUNW), metabolically unhealthy overweight (MUOW), and MUO. A multistate Markov model was used to analyse all possible transitions among these phenotypes and assess the effects of demographic and blood indicators on the transitions.ResultsThe transition intensity from MUNW to MHNW was the highest (0.64), followed by the transition from MHO to MUO (0.56). The greatest sojourn time appeared in the MHNW state (3.84 years), followed by the MUO state (2.34 years), and the shortest sojourn time appeared in the MHO state (1.16 years). Transition intensities for metabolic improvement gradually decreased with BMI level as follows: 0.64 for MUNW to MHNW, 0.44 for MUOW to MHNW, and 0.27 for MUO to MHO; however, transition intensities for metabolic deterioration, including MHNW to MUNW, MHOW to MUOW, and MHO to MUO, were 0.15, 0.38, and 0.56, respectively. In the middle-aged male group, elevated alanine aminotransferase (ALT), aspartate aminotransferase (AST), and uric acid (UA) increased the risk of deterioration in weight and metabolic status and decreased the possibility of improvement.ConclusionMaintaining a normal and stable BMI is important for metabolic health. More attention should be given to males and elderly people to prevent their progression to an unhealthy metabolic and/or weight status. MHO is the most unstable phenotype and is prone to convert to the MUO state, and individuals with abnormal ALT, AST and UA are at an increased risk of transitioning to an unhealthy weight and/or metabolic status; therefore, we should be alert to abnormal indicators and MHO. Intervention measures should be taken early to maintain healthy weight and metabolic status.</p

Ultrastrong, Foldable, and Highly Conductive Carbon Nanotube Film

Preparation of strong, flexible, and multifunctional carbon-based films has attracted considerable interest not only in fundamental research areas but also for industrial applications. We report a binder-free, ultrastrong, and foldable carbon nanotube (CNT) film using aligned few-walled nanotube sheets drawn from spinnable nanotube arrays. The film exhibits tensile strengths up to ∼2 GPa and a Young’s modulus up to ∼90 GPa, which is markedly superior to other types of carbon-based films reported, including commercial graphite foils, buckypapers, and graphene-related papers. The film can bear severe bending (even being folded) and shows good structure integrity and negligible change in electric conductivity. The unique structure of the CNT film (good nanotube alignment, high packing density) provides the film with direct and efficient transport paths for electricity. As a flexible charge collector, it favors a magnesium oxide coating to exhibit high charge/discharge rate stability and an excellent electrochemical capacitance close to its theoretical value

Effective Optimization of Antibody Affinity by Phage Display Integrated with High-Throughput DNA Synthesis and Sequencing Technologies

<div><p>Phage display technology has been widely used for antibody affinity maturation for decades. The limited library sequence diversity together with excessive redundancy and labour-consuming procedure for candidate identification are two major obstacles to widespread adoption of this technology. We hereby describe a novel library generation and screening approach to address the problems. The approach started with the targeted diversification of multiple complementarity determining regions (CDRs) of a humanized anti-ErbB2 antibody, HuA21, with a small perturbation mutagenesis strategy. A combination of three degenerate codons, NWG, NWC, and NSG, were chosen for amino acid saturation mutagenesis without introducing cysteine and stop residues. In total, 7,749 degenerate oligonucleotides were synthesized on two microchips and released to construct five single-chain antibody fragment (scFv) gene libraries with 4 x 10⁶ DNA sequences. Deep sequencing of the unselected and selected phage libraries using the Illumina platform allowed for an in-depth evaluation of the enrichment landscapes in CDR sequences and amino acid substitutions. Potent candidates were identified according to their high frequencies using NGS analysis, by-passing the need for the primary screening of target-binding clones. Furthermore, a subsequent library by recombination of the 10 most abundant variants from four CDRs was constructed and screened, and a mutant with 158-fold increased affinity (<i>K_d</i> = 25.5 pM) was obtained. These results suggest the potential application of the developed methodology for optimizing the binding properties of other antibodies and biomolecules.</p></div

Frequency distribution of CDR sequences.

<p>Data analysis was performed for five individual CDRs from 13.9 and 4.5 million qualified sequences in the unselected (BS) and selected (AS) libraries, respectively. Total counts of all variants representing that particular frequency group were calculated.</p

Summary of the NGS data in the unselected and selected libraries.

<p>a: Number of raw sequences that pass the quality-filtering steps.</p><p>b: Number of sequences, variants, or peptides mutants that have the expected CDR lengths and accurate in-frame reading.</p><p>c: Number of sequences, variants, or peptides mutants that perfectly match the CDR diversification strategy and degenerate codon design.</p><p>d: Number of unique DNA sequences.</p><p>e: Number of DNA sequences occurring only one time.</p><p>Summary of the NGS data in the unselected and selected libraries.</p

Characterization of library CDR sequences.

<p>Data analysis was performed for the five individual CDRs from 13.9 million qualified sequences in the unselected libraries. (A) Sequence abundance analysis. Each point represents the total number of variants with a specific copy number. Only variants with ≤10 copies are shown. (B) Length distribution analysis. Each point represents the total number of variants of a specific length. Only variants with ≤5 nucleotide deletions or insertions are shown.</p

Affinity gains by target-binding screening and NGS approaches.

<p>The scFv-Fc mutants were expressed in Expi293 cells to determine their apparent binding affinities to ErbB2 by screening ELISA. The fold affinity enhancement was calculated as the EC50 ratio of wild-type HuA21/mutant. The numbers of mutants from different CDR libraries are indicated.</p

Binding kinetics of the combinatorial library mutants.

<p>Binding kinetics of the combinatorial library mutants.</p

Statistics of the SPM libraries for CDR diversification.

<p>a: Underlined resides are selected as candidate positions.</p><p>b: The number of sub-libraries was calculated using the following formula: C³/_n, where n is the number of candidate positions in each CDR.</p><p>Statistics of the SPM libraries for CDR diversification.</p

Amino acid composition at different CDR positions.

<p>(A) Map of the theoretical amino acid composition using NWG, NWC and NSG degenerate codons according to the SPM strategy. (B) Map of the observed amino acid composition from 10.3 million functional sequences in the unselected libraries. Forty diversified positions and four non-diversified positions are shown. Z represents a stop codon.</p