Distinguish MnO2/Mn2+ Conversion/ Zn2+ Intercalation/ H+ Conversion Chemistries at Different Potentials in Aqueous Zn||MnO2 Batteries.

The rechargeable aqueous Zn||MnO2 chemistry has been extensively explored, but its electrochemical reaction mechanisms, especially in the context of MnO2/Mn2+ conversion and Zn2+/H+ intercalation chemistry, remain not fully understood. Here, we designed an amphiphilic hydrogel electrolyte, which distinguished the MnO2/Mn2+ conversion, Zn2+ intercalation, and H+ intercalation and conversion processes at three distinct discharge plateaus of an aqueous Zn||MnO2 battery. The amphiphilic hydrogel electrolyte is featured with an extended electrochemical stability window up to 3.0 V, high ionic conductivity, Zn2+-selective ion tunnels, and hydrophobic associations with cathode materials. This specifically designed electrolyte allows the MnO2/Mn2+ conversion reaction at a discharge plateau of 1.75 V. More interesting, the discharge plateaus of ~1.33 V, previously assigned as the co-intercalation of Zn2+ and H+ ions in the MnO2 cathode, are specified as the exclusive intercalation of Zn2+ ions, leading to an ultra-flat voltage plateau. Furthermore, with a distinct three-step electrochemical energy storage process, a high areal capacity of 1.8 mAh cm-2 and high specific energy of 0.858 Wh cm-2, even at a low MnO2 loading mass of 0.5 mg cm-2 are achieved. To our knowledge, this is the first report to fully distinguish different mechanisms at different potentials in aqueous Zn||MnO2 batteries.

Ssc-miR-141 Attenuates Hypoxia-Induced Alveolar Type II Epithelial Cell Injury in Tibetan Pigs by Targeting PDCD4.

The Tibetan pig is an endemic economic animal in the plateau region of China, and has a unique adaptation mechanism to the plateau hypoxic environment. Research into microRNAs (miRNAs) involved in the mechanism underlying hypoxia adaptation of Tibetan pig is very limited. Therefore, we isolated alveolar type II epithelial (ATII) cells from the lungs of the Tibetan pig, cultured them in normoxia/hypoxia (21% O2; 2% O2) for 48 h, and performed high-throughput sequencing analysis. We identified a hypoxic stress-related ssc-miR-141 and predicted its target genes. The target genes of ssc-miR-141 were mainly enriched in mitogen-activated protein kinase (MAPK), autophagy-animal, and Ras signaling pathways. Further, we confirmed that PDCD4 may serve as the target gene of ssc-miR-141. Real-time quantitative polymerase chain reaction (RT-qPCR) analysis was performed to confirm the expression levels of ssc-miR-141 and PDCD4, and a dual-luciferase gene reporter system was used to verify the targeted linkage of ssc-miR-141 to PDCD4. The results showed that the expression level of ssc-miR-141 in the hypoxia group was higher than that in the normoxia group, while the expression level of PDCD4 tended to show the opposite trend and significantly decreased under hypoxia. These findings suggest that ssc-miR-141 is associated with hypoxia adaptation and provide a new insight into the role of miRNAs from ATII cells of Tibetan pig in hypoxia adaptation.

Characterization of circRNA-miRNA-mRNA networks regulating oxygen utilization in type II alveolar epithelial cells of Tibetan pigs.

Understanding the signaling pathway regulatory mechanisms in type II alveolar epithelial (ATII) cells, the progenitor cells responsible for proliferating and regenerating type I alveolar epithelial (ATI) and ATII cells, in Tibetan pigs is beneficial for exploring methods of preventing and repairing cellular damage during hypoxia. We simulated a hypoxic environment (2% O2) for culture ATII cells of Tibetan pigs and Landrace pigs, with cells cultured under normoxic conditions (21% O2) as a control group, and performed integrated analysis of circular RNA (circRNA)-microRNA (miRNA)-messenger RNA (mRNA) regulatory axes by whole-transcriptome sequencing. Functional enrichment analysis indicated that the source genes of the differential expressed circRNAs (DEcircRNAs) were primarily involved in cell proliferation, cellular processes, and cell killing. A series of DEcircRNAs were derived from inhibitors of apoptosis proteins and led to a key autonomous effect as modulators of cell repair in Tibetan pigs under hypoxia. The significant higher expression of COL5A1 in TL groups may inhibited apoptosis of ATII cells in Tibetan pigs under lower oxygen concentration, and may lead their better survive in the hypoxia environment. In addition, a competing endogenous RNA (ceRNA) network of functional interactions was constructed that included novel_circ_000898-ssc-miR-199a-5p-CAV1 and novel_circ_000898-ssc-miR-378-BMP2, based on the node genes ssc-miR-199a-5p and ssc-miR-378, which may regulate multiple miRNAs and mRNAs that mediate endoplasmic reticulum (ER) stress-induced apoptosis and inflammation and attenuate hypoxia-induced injury in ATII cells under hypoxic conditions. These results broaden our knowledge of circRNAs, miRNAs, and mRNAs associated with hypoxia and provide new insights into the hypoxic response of ATII cells in Tibetan pigs.

Alternative splicing signature of alveolar type II epithelial cells of Tibetan pigs under hypoxia-induced.

Alternative splicing (AS) allows the generation of multiple transcript variants from a single gene and affects biological processes by generating protein diversity in organisms. In total, 41,642 AS events corresponding to 9,924 genes were identified, and SE is the most abundant alternatively spliced type. The analysis of functional categories demonstrates that alternatively spliced differentially expressed genes (DEGs) were enriched in the MAPK signaling pathway and hypoxia-inducible factor 1 (HIF-1) signaling pathway. Proteoglycans in cancer between the normoxic (21% O2, TN and LN) and hypoxic (2% O2, TL and LL) groups, such as SLC2A1, HK1, HK2, ENO3, and PFKFB3, have the potential to rapidly proliferate alveolar type II epithelial (ATII) cells by increasing the intracellular levels of glucose and quickly divert to anabolic pathways by glycolysis intermediates under hypoxia. ACADL, EHHADH, and CPT1A undergo one or two AS types with different frequencies in ATII cells between TN and TL groups (excluding alternatively spliced DEGs shared between normoxic and hypoxic groups), and a constant supply of lipids might be obtained either from the circulation or de novo synthesis for better growth of ATII cells under hypoxia condition. MCM7 and MCM3 undergo different AS types between LN and LL groups (excluding alternatively spliced DEGs shared between normoxic and hypoxic groups), which may bind to the amino-terminal PER-SIM-ARNT domain and the carboxyl terminus of HIF-1α to maintain their stability. Overall, AS and expression levels of candidate mRNAs between Tibetan pigs and Landrace pigs revealed by RNA-seq suggest their potential involvement in the ATII cells grown under hypoxia conditions.

Integrative Analysis of the lncRNA-Associated ceRNA Regulatory Network Response to Hypoxia in Alveolar Type II Epithelial Cells of Tibetan Pigs.

The function of alveolar type II epithelial (ATII) cells is severely hampered by oxygen deficiency, and understanding the regulatory mechanisms controlling responses to hypoxia may assist in relieving injury induced by hypoxia. In this study, we cultured ATII cells from Tibetan pigs and Landrace pigs under hypoxic and normoxic environments to screen for differentially expressed (DE) lncRNAs, DEmiRNAs, and construct their associated ceRNA regulatory networks in response to hypoxia. Enrichment analysis revealed that target genes of DElncRNAs of Tibetan pigs and Landrace pig between the normoxic (TN, LN) and hypoxic (TL, LL) groups significantly enriched in the proteoglycans in cancer, renal cell carcinoma, and erbB signaling pathways, while the target genes of DEmiRNAs were significantly enriched in the axon guidance, focal adhesion, and mitogen-activated protein kinase (MAPK) signaling pathways. Hypoxia induction was shown to potentially promote apoptosis by activating the focal adhesion/PI3K-Akt/glycolysis pathway. The ssc-miR-20b/MSTRG.57127.1/ssc-miR-7-5p axis potentially played a vital role in alleviating hypoxic injury by regulating ATII cell autophagy under normoxic and hypoxic conditions. MSTRG.14861.4-miR-11971-z-CCDC12, the most affected axis, regulated numerous RNAs and may thus regulate ATII cell growth in Tibetan pigs under hypoxic conditions. The ACTA1/ssc-miR-30c-3p/MSTRG.23871.1 axis is key for limiting ATII cell injury and improving dysfunction and fibrosis mediated by oxidative stress in Landrace pigs. Our findings provide a deeper understanding of the lncRNA/miRNA/mRNA regulatory mechanisms of Tibetan pigs under hypoxic conditions.

Transcriptome and Metabolome Integration Provides New Insights Into the Regulatory Networks of Tibetan Pig Alveolar Type II Epithelial Cells in Response to Hypoxia.

Tibetan pigs show a widespread distribution in plateau environments and exhibit striking physiological and phenotypic differences from others pigs for adaptation to hypoxic conditions. However, the regulation of mRNAs and metabolites as well as their functions in the alveolar type II epithelial (ATII) cells of Tibetan pigs remain undefined. Herein, we carried out integrated metabolomic and transcriptomic profiling of ATII cells between Tibetan pigs and Landrace pigs across environments with different oxygen levels to delineate their signature pathways. We observed that the differentially accumulated metabolites (DAMs) and differentially expressed genes (DEGs) profiles displayed marked synergy of hypoxia-related signature pathways in either Tibetan pigs or Landrace pigs. A total of 1,470 DEGs shared between normoxic (TN, ATII cells of Tibetan pigs were cultured under 21% O2; LN, ATII cells of Landrace pigs were cultured under 21% O2) and hypoxic (TL, ATII cells of Tibetan pigs were cultured under 2% O2; LL, ATII cells of Landrace pigs were cultured under 2% O2) groups and 240 DAMs were identified. Functional enrichment assessment indicated that the hypoxia-related genes and metabolites were primarily involved in glycolysis and aldosterone synthesis and secretion. We subsequently constructed an interaction network of mRNAs and metabolites related to hypoxia, such as guanosine-3', 5'-cyclic monophosphate, Gly-Tyr, and phenylacetylglycine. These results indicated that mitogen-activated protein kinase (MAPK) signaling, aldosterone synthesis and secretion, and differences in the regulation of MCM and adenosine may play vital roles in the better adaptation of Tibetan pigs to hypoxic environments relative to Landrace pigs. This work provides a new perspective and enhances our understanding of mRNAs and metabolites that are activated in response to hypoxia in the ATII cells of Tibetan pigs.

Post-transcriptional regulation through alternative splicing in the lungs of Tibetan pigs under hypoxia.

In multicellular organisms, alternative splicing (AS) is central to the regulation of multiple biological processes. To further elucidate the adaptive strategy of AS in the lungs of Tibetan pigs in response to hypoxia, we identified and analyzed five basic AS types and 59,930 AS events in 18,179 genes. We found that approximately 65.10% of the total expressed genes underwent AS in the lungs of Tibetan pigs at a high altitude (TH). The frequencies of AS events were similar among the different groups (5.06-5.30 events in each gene on average). Skipped exons (SEs) were the predominant type of AS event, followed by mutually exclusive exons (MXEs), alternative 3' splice sites (A3SSs) and alternative 5' splice sites (A5SSs). Retained introns (RIs), the remaining type of AS event, showed lower frequencies. Further comparison analysis of differentially expressed genes (DEGs) and differentially spliced genes (DSGs) identified 2,209 differential splicing events in the above 18,000 expressed genes, including 918 increased and 1,291 decreased splicing events between the TH and Tibetan pigs at a low altitude (TL) groups. We identified 227 hypoxia-related genes involved in lung development that were differentially regulated through AS. Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis clearly identified many DEGs and DSGs at high or low altitude. Seven pathways in the top 20 enriched KEGG terms overlapped for the DEGs and DSGs, including the chemokine signaling pathway, B cell receptor signaling pathway, and cytokine-cytokine receptor interaction, which exert many immunoregulatory and inflammatory actions critical to the lung under hypoxia. Twelve pathways overlapped in hypoxic DEGs and DSGs and included antigen processing, presentation and biosynthesis. ​GO analysis of the DEGs and DSGs among the four groups showed that numerous GO terms were enriched in the biological category, and the proportion of genes with downregulated expression was greater among 227 hypoxic genes than that of all genes. The results suggest that AS plays an essential role in the regulation of gene expression during hypoxia and that numerous genes involved in lung development are differentially regulated through AS.

The Expression Regulatory Network in the Lung Tissue of Tibetan Pigs Provides Insight Into Hypoxia-Sensitive Pathways in High-Altitude Hypoxia.

To adapt to a low-oxygen environment, Tibetan pigs have developed a series of unique characteristics and can transport oxygen more effectively; however, the regulation of the associated processes in high-altitude animals remains elusive. We performed mRNA-seq and miRNA-seq, and we constructed coexpression regulatory networks of the lung tissues of Tibetan and Landrace pigs. HBB, AGT, COL1A2, and EPHX1 were identified as major regulators of hypoxia-induced genes that regulate blood pressure and circulation, and they were enriched in pathways related to signal transduction and angiogenesis, such as HIF-1, PI3K-Akt, mTOR, and AMPK. HBB may promote the combination of hemoglobin and oxygen as well as angiogenesis for high-altitude adaptation in Tibetan pigs. The expression of MMP2 showed a similar tendency of alveolar septum thickness among the four groups. These results indicated that MMP2 activity may lead to widening of the alveolar wall and septum, alveolar structure damage, and collapse of alveolar space with remarkable fibrosis. These findings provide a perspective on hypoxia-adaptive genes in the lungs in addition to insights into potential candidate genes in Tibetan pigs for further research in the field of high-altitude adaptation.

Influence of the second phase on protein adsorption on biodegradable Mg alloys' surfaces: Comparative experimental and molecular dynamics simulation studies.

The effect of the second phase on the mechanical properties and corrosion resistance of Mg alloys has been systematically studied. However, there is limited information on the effect of the second phase on protein adsorption behavior. In the present study, the effect of the second phase on protein adsorption on the surfaces of biodegradable Mg alloys was investigated using experimental methods and molecular dynamics (MD) simulations. The experimental results showed that the effect of the second phase on fibrinogen adsorption was type-dependent. Fibrinogen preferentially adsorbed on Y-, Ce-, or Nd-involved second phases, while the second phase containing Zn inhibited its adsorption. MD simulations revealed the mechanism of the second phase that influenced protein adsorption in terms of charge distribution, surface-protein interaction energy, and water molecule distribution. Our studies proposed a deep understanding of the design of Mg-based biomaterials with superior biocompatibility. STATEMENT OF SIGNIFICANCE: Mechanical properties, uniform degradation, and biocompatibility must be considered while designing biomedical Mg alloys. To improve the mechanical properties and corrosion resistance of Mg alloys, the second phase is usually required. However, the effects of the second phase on biocompatibility of Mg alloys have been rarely reported. Here, the influence of the second phase on protein adsorption was experimentally studied by designing Mg alloys with different types of second phase. The first principle calculation and MD simulation were used to reveal the mechanism by which the second phase influences protein adsorption. This work could be used to better elucidate the protein adsorption mechanisms and design principles to improve the biocompatibility of Mg alloys.

25-Methoxylprotopanaxadiol derivatives and their anti-proliferative activities.

(20R)-25-Methoxyl-dammarane-3ß,12ß,20-triol (25-OCH3-PPD) is a dammarane-type sapogenin showing anti-proliferative potential. In our study, two series of analogs substituted at the C-3 or C-3 and C-12 positions with fatty acids were prepared conveniently. The cytotoxic activity of these compounds was evaluated using four different human tumor cell lines (A549, Hela, HT-29 and MCF-7) and a normal cell line (IOSE144). As compared with 25-OCH3-PPD, compounds 1a, 1b, 2a and 2b showed better anti-proliferative activities against all tumor cell lines and all the derivatives, with low toxicities in the normal cell line. Compound 1a (C-3 monoformiate) exhibited the strongest activity with the IC50 value of 5.2 µM towards HT29 cells. The results indicated that the difference in the substituents may affect the anti-proliferative activity of the compounds. The longer the side chain of 25-OCH3-PPD is, the lower the anti-proliferative activity would be. This information may be useful for evaluating the structure-activity relationship of other dammarane-type sapogenins and for development of novel antineoplastic agents.

Synthesis and anti-tumor evaluation of novel 25-hydroxyprotopanaxadiol analogs incorporating natural amino acids.

In the current study, derivatives of 25-hydroxyprotopanaxadiol (25-OH-PPD) were prepared and their in vitro anti-tumor activities were tested on six different human tumor cell lines by standard MTT assay. The results showed that combining an ester group combined with the presence of an amino acid moiety led to a 10-fold improved anti-tumor activity. Compound 1c exhibited the best anti-tumor activity in the in vitro assays. Compounds 2c, 3c, 4c, 5c, 6c and 8b showed better anti-tumor activities compared to the parent compound 25-OH-PPD. The current results may provide useful data for researching and developing new anti-cancer agents.

Semi-synthesis and anti-tumor evaluation of novel 25-hydroxyprotopanaxadiol derivatives.

30 novel compounds have been synthesized from 25-hydroxyprotopanaxadiol (25-OH-PPD) and their in vitro anti-tumor activities were tested on three cancer cell lines and one normal cell line (IOSE144) by standard MTT assay. The results showed that compound 27 exhibited the best anti-tumor activity in the in vitro assays. Compounds 1, 2, 16, 17, 18, 27, 28 and 29 have better anti-tumor activities against MCF-7 and A549 cancer cell lines than 25-OH-PPD, together with low toxicity in the normal cell. The results may provide useful data for researching and developing new antitumor agents.