Integrated transcriptomic and metabolomic analyses elucidate the mechanism of flavonoid biosynthesis in the regulation of mulberry seed germination under salt stress.

Seed propagation is the main method of mulberry expansion in China, an important economic forest species. However, seed germination is the most sensitive stage to various abiotic stresses, especially salinity stress. To reveal the molecular regulatory mechanism of mulberry seed germination under salt stress, flavonoid metabolomics and transcriptomics analyses were performed on mulberry seeds germinated under 50 and 100 mmol/L NaCl stress. Analysis of the flavonoid metabolome revealed that a total of 145 differential flavonoid metabolites (DFMs) were classified into 9 groups, 40 flavonols, 32 flavones, 16 chalcones and 14 flavanones. Among them, 61.4% (89) of the DFMs accumulated continuously with increasing salt concentration, reaching the highest level at a 100 mmol/L salt concentration; these DFMs included quercetin-3-O-glucoside (isoquercitrin), kaempferol (3,5,7,4'-tetrahydroxyflavone), quercetin-7-O-glucoside, taxifolin (dihydroquercetin) and apigenin (4',5,7-trihydroxyflavone), indicating that these flavonoids may be key metabolites involved in the response to salt stress. Transcriptional analysis identified a total of 3055 differentially expressed genes (DEGs), most of which were enriched in flavonoid biosynthesis (ko00941), phenylpropanoid biosynthesis (ko00940) and biosynthesis of secondary metabolites (ko01110). Combined analysis of flavonoid metabolomic and transcriptomic data indicated that phenylalanine ammonia-lyase (PAL), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), flavonol synthase (FLS), bifunctional dihydroflavonol 4-reductase/flavanone 4-reductase (DFR) and anthocyanidin reductase (ANR) were the key genes involved in flavonoid accumulation during mulberry seed germination under 50 and 100 mmol/L NaCl stress. In addition, three transcription factors, MYB, bHLH and NAC, were involved in the regulation of flavonoid accumulation under salt stress. The results of quantitative real-time PCR (qRT‒PCR) validation showed that the expression levels of 11 DEGs, including 7 genes involved in flavonoid biosynthesis, under different salt concentrations were consistent with the transcriptomic data, and parallel reaction monitoring (PRM) results showed that the expression levels of 6 key enzymes (proteins) involved in flavonoid synthesis were consistent with the accumulation of flavonoids. This study provides a new perspective for investigating the regulatory role of flavonoid biosynthesis in the regulation of mulberry seed germination under salt stress at different concentrations.

CircGUCY2C regulates cofilin 1 by sponging miR-425-3p to promote the proliferation of porcine skeletal muscle satellite cells.

Circular ribonucleic acids (or circRNAs) are an emerging class of endogenous noncoding RNAs that are involved in physiological and pathological processes. Increasing evidence suggests that circRNAs play an important regulatory role in skeletal muscle development and meat quality regulation. In this study, it was found that circGUCY2C exhibits a high expression level in the longissimus dorsi muscle. It shows resistance to RNase R and additionally promotes the mRNA expression of cyclin-dependent kinase 2 (CDK2) and proliferating cell nuclear antigen (PCNA). Specifically, it was observed that the overexpression of circGUCY2C could promote the transition of porcine skeletal muscle satellite cells into the S and G2 phases of the cell cycle and that it regulates the proliferation of porcine skeletal muscle satellite cells. In contrast, miR-425-3p plays the opposite role and has an inhibitory effect on the proliferation of porcine skeletal muscle satellite cells. MiR-425-3p has been described as a target of circGUCY2C; consequently, the depletion of miR-425-3p promoted the proliferation of porcine skeletal muscle satellite cells. CFL1 (cofilin 1) is a target of miR-425-3p, and circGUCY2C upregulated CFL1 expression by inhibiting miR-425-3p. Collectively, our research outcomes demonstrate that circGUCY2C significantly influences the proliferation of porcine skeletal muscle satellite cells by selectively targeting the miR-425-3p-CFL1 axis, and our work partially clarified the role of circGUCY2C in porcine skeletal muscle satellite cells. Thus, the study provides new insight into the function of circGUCY2C and adds to the knowledge of the post-transcriptional regulation of pork quality.

Genome-Wide Analysis of MAMSTR Transcription Factor-Binding Sites via ChIP-Seq in Porcine Skeletal Muscle Fibroblasts.

Myocyte enhancer factor-2-activating motif and SAP domain-containing transcriptional regulator (MAMSTR) regulates its downstream through binding in its promoter regions. However, its molecular mechanism, particularly the DNA-binding sites, and coregulatory genes are quite unexplored. Therefore, to identify the genome-wide binding sites of the MAMSTR transcription factors and their coregulatory genes, chromatin immunoprecipitation sequencing was carried out. The results showed that MAMSTR was associated with 1506 peaks, which were annotated as 962 different genes. Most of these genes were involved in transcriptional regulation, metabolic pathways, and cell development and differentiation, such as AMPK signaling pathway, TGF-beta signaling pathway, transcription coactivator activity, transcription coactivator binding, adipocytokine signaling pathway, fat digestion and absorption, skeletal muscle fiber development, and skeletal muscle cell differentiation. Lastly, the expression levels and transcriptional activities of PID1, VTI1B, PRKAG1, ACSS2, and SLC28A3 were screened and verified via functional markers and analysis. Overall, this study has increased our understanding of the regulatory mechanism of MAMSTR during skeletal muscle fibroblast development and provided a reference for analyzing muscle development mechanisms.

Transcriptome-wide analysis of RNA m6A methylation regulation of muscle development in Queshan Black pigs.

BACKGROUND: N6-methyladenosine (m6A) refers to the methylation modification of N6 position of RNA adenine, a dynamic reversible RNA epigenetic modification that plays an important regulatory role in a variety of life processes. In this study, we used MeRIP-Seq and RNA-Seq of the longissimus dorsi (LD) muscle of adult (QA) and newborn (QN) Queshan Black pigs to screen key genes with m6A modification involved in muscle growth by bioinformatics analysis. RESULTS: A total of 23,445 and 25,465 m6A peaks were found in the whole genomes of QA and QN, respectively. Among them, 613 methylation peaks were significantly different (DMPs) and 579 genes were defined as differentially methylated genes (DMGs). Compared with the QN group, there were 1,874 significantly differentially expressed genes (DEGs) in QA group, including 620 up-regulated and 1,254 down-regulated genes. In order to investigate the relationship between m6A and mRNA expression in the muscle of Queshan Black pigs at different periods, a combined analysis of MeRIP-Seq and RNA-Seq showed that 88 genes were significantly different at both levels. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes results showed that DEGs and DMGs were mainly involved in skeletal muscle tissue development, FoxO signaling pathway, MAPK signaling pathway, insulin signaling pathway, PI3K-Akt signaling pathway, and Wnt signaling pathway. Four DEGs (IGF1R, CCND2, MYOD1 and FOS) and four DMGs (CCND2, PHKB, BIN1 and FUT2), which are closely related to skeletal muscle development, were selected as candidate genes for verification, and the results were consistent with the sequencing results, which indicated the reliability of the sequencing results. CONCLUSIONS: These results lay the foundation for understanding the specific regulatory mechanisms of growth in Queshan Black pigs, and provide theoretical references for further research on the role of m6A in muscle development and breed optimization selection.

Effect of miR-149-5p on intramuscular fat deposition in pigs based on metabolomics and transcriptomics.

As one of the important traits in pig production, meat quality has important research significance and value. Intramuscular fat (IMF) content is one of the most important factors affecting pork quality. Many experimental studies have shown that IMF content is closely related to the flavor, tenderness, and juiciness of pork. Therefore, it is of great significance to study the mechanism of porcine IMF deposition. Previous research indicated that miR-149-5p promoted the proliferation of porcine intramuscular (IM) preadipocytes and decreased their ability to differentiate, albeit the exact mechanism of action is unknown. In vitro, foreign pigs showed increased miR-149-5p expression and reduced fat deposition when compared to Queshan Black pigs. This study conducted metabolomics and transcriptomics analyses of porcine IM preadipocytes overexpressing miR-149-5p to verify their effects on lipid formation. According to metabolomics analysis, the overexpression of miR-149-5p has significantly altered the lipid, organic acid, and organic oxygen metabolites of porcine IM preadipocytes. Specially speaking, it has changed 115 metabolites, including 105 up-regulated and 10 down-regulated ones, as well as the composition of lipid, organic acid, and organic oxygen metabolism-related metabolites. RNA-seq analysis showed that overexpression of miR-149-5p significantly altered 857 genes, of which 442 were up-regulated, and 415 were down-regulated, with enrichment to MAPK, IL-17, PI3K-Akt, and ErbB signaling pathways. We found that overexpression of miR-149-5p inhibited adipogenic differentiation by changing cAMP signaling pathway in porcine IM preadipocytes. In addition, the overexpression of miR-149-5p may affect the transport of Cu2+ by targeting ATP7A and inhibiting adipogenic differentiation. These findings elucidate the regulatory function of miR-149-5p in porcine IM preadipocytes, which may be a key target for controlling pork quality.

Physiological and Proteomic Analysis of Seed Germination under Salt Stress in Mulberry.

BACKGROUND: Salinity is the main abiotic stress that affects seed germination, plant growth and crop production. Plant growth begins with seed germination, which is closely linked to crop development and final yields. Morus alba L. is a well-known saline-alkaline tree with economic value in China, and the most prominent method of expanding mulberry tree populations is seed propagation. Understanding the molecular mechanism of Morus alba L. salt tolerance is crucial for identifying salt-tolerant proteins in seed germination. Here, we explored the response mechanism of mulberry seed germination to salt stress at physiological and protein omics levels. METHODS: Tandem mass tag (TMT)-based proteomic profiling of Morus alba L. seeds germinated under 50 mM and 100 mM NaCl treatment for 14 days was performed, and the proteomic findings were validated through parallel reaction monitoring (PRM). RESULTS: Physiological data showed that salt stress inhibited the germination rate and radicle length of mulberry seeds, decreased the malondialdehyde (MDA) content and significantly increased superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activities. Then, a TMT marker technique was used to analyze the protein groups in mulberry seeds with two salt treatment stages, and 76,544 unique peptides were detected. After removing duplicate proteins, 7717 proteins were identified according to TMT data, and 143 (50 mM NaCl) and 540 (100 mM NaCl) differentially abundant proteins (DAPs) were screened out. Compared with the control, in the 50 mM NaCl solution, 61 and 82 DAPs were upregulated and downregulated, respectively, and in the 100 mM NaCl solution, 222 and 318 DAPs were upregulated and downregulated, respectively. Furthermore, 113 DAPs were copresent in the 50 mM and 100 mM NaCl treatments, of which 43 were upregulated and 70 were downregulated. Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed that the DAPs induced by salt stress during mulberry seed germination were mainly involved in photosynthesis, carotenoid biosynthesis and phytohormone signaling. Finally, PRM verified five differentially expressed proteins, which demonstrated the reliability of TMT in analyzing protein groups. CONCLUSIONS: Our research provides valuable insights to further study the overall mechanism of salt stress responses and salt tolerance of mulberry and other plants.

Identification and Functional Prediction of Long Non-Coding RNA in Longissimus Dorsi Muscle of Queshan Black and Large White Pigs.

Long non-coding RNA (lncRNA) participates in the regulation of various biological processes, but its function and characteristics in intramuscular fat (IMF) deposition in different breeds of pigs have not been fully understood. IMF content is one of the important factors affecting pork quality. In the present study, the differentially expressed lncRNAs (DE lncRNAs) and their target genes were screened by comparing Queshan Black (QS) and Large White (LW) pigs based on RNA-seq. The results displayed 55 DE lncRNAs between QS and LW, 29 upregulated and 26 downregulated, with 172 co-located target genes, and 6203 co-expressed target genes. The results of GO and KEGG analysis showed that the target genes of DE lncRNAs were involved in multiple pathways related to lipogenesis and lipid metabolism, such as the lipid biosynthetic process, protein phosphorylation, activation of MAPK activity, and the Jak-STAT signaling pathway. By constructing regulatory networks, lincRNA-ZFP42-ACTC1, lincRNA-AMY2-STAT1, and/or lincRNA-AMY2/miR-204/STAT1 were sieved, and the results indicate that lncRNA could participate in IMF deposition through direct regulation or ceRNA. These findings provide a basis for analyzing the molecular mechanism of IMF deposition in pigs and lay a foundation for developing and utilizing high-quality resources of local pig breeds.

Survival, symptoms and hospitalization of older patients with advanced chronic kidney disease managed without dialysis.

BACKGROUND: Shared decision-making (SDM) is important when considering whether an older patient with advanced chronic kidney disease (CKD) should be managed with dialysis or conservative kidney management (CKM). Physicians may find these conversations difficult because of the relative paucity of data on patients managed without dialysis. METHODS: This prospective observational study was conducted in a unit supported by a multidisciplinary Kidney Supportive Care (KSC) programme, in a cohort of 510 patients (280 CKM and 230 dialysis) ≥65 years of age with CKD stages 4 and 5. Survival was evaluated using logistic regression and Cox proportional hazards models. Linear mixed models were utilized to assess symptoms over time. RESULTS: CKM patients were older (mean 84 versus 74 years; P < .001) and almost 2-fold more likely to have three or more comorbidities (P < .001). The median survival of CKM patients was lower compared with dialysis from all time points: 14 months [interquartile range (IQR) 6-32] versus 53 (IQR 28-103) from decision of treatment modality or dialysis start date (P < .001); 15 months (IQR 7-34) versus 64 (IQR 30-103) from the time the estimated glomerular filtration rate (eGFR) was ≤15 mL/min/1.73 m2 (P < .001); and 8 months (IQR 3-18) versus 49 (19-101) from eGFR ≤10 mL/min/1.73 m2. A total of 59% of CKM patients reported an improvement in symptoms by their third KSC clinic visit (P < .001). The rate of unplanned hospitalization was 2-fold higher in the dialysis cohort. CONCLUSIONS: CKM patients survive a median of 14 months from the time of modality choice and have a lower rate of hospitalization than dialysis patients. Although the symptom burden in advanced CKD is high, most elderly CKM patients managed through an integrated KSC programme and can achieve improvement in their symptoms over time. These data might help with SDM.

Treatment decision-making and care among older adults with kidney failure: protocol for a multicentre, prospective observational cohort study with nested substudies and linked qualitative research (the Elderly Advanced CKD Programme).

INTRODUCTION: Shared treatment decision-making and planning of care are fundamental in advanced chronic kidney disease (CKD) management. There are limited data on several key outcomes for the elderly population including survival, quality of life, symptom burden, changes in physical functioning and experienced burden of healthcare. Patients, caregivers and clinicians consequently face significant uncertainty when making life-impacting treatment decisions. The Elderly Advanced CKD Programme includes quantitative and qualitative studies to better address challenges in treatment decision-making and planning of care among this increasingly prevalent elderly cohort. METHODS AND ANALYSIS: The primary component is OUTcomes of Older patients with Kidney failure (OUTLOOK), a multicentre prospective observational cohort study that will enrol 800 patients ≥75 years with kidney failure (estimated glomerular filtration rate ≤15 mL/min/1.73 m2) across a minimum of six sites in Australia. Patients entered are in the decision-making phase or have recently made a decision on preferred treatment (dialysis, conservative kidney management or undecided). Patients will be prospectively followed until death or a maximum of 4 years, with the primary outcome being survival. Secondary outcomes are receipt of short-term acute dialysis, receipt of long-term maintenance dialysis, changes in biochemistry and end-of-life care characteristics. Data will be used to formulate a risk prediction tool applicable for use in the decision-making phase. The nested substudies Treatment modalities for the InfirM ElderLY with end stage kidney disease (TIMELY) and Caregivers of The InfirM ElderLY with end stage kidney disease (Co-TIMELY) will longitudinally assess quality of life, symptom burden and caregiver burden among 150 patients and 100 caregivers, respectively. CONsumer views of Treatment options for Elderly patieNts with kiDney failure (CONTEND) is an additional qualitative study that will enrol a minimum of 20 patients and 20 caregivers to explore experiences of treatment decision-making and care. ETHICS AND DISSEMINATION: Ethics approval was obtained through Sydney Local Health District Human Research Ethics Committee (2019/ETH07718, 2020/ETH02226, 2021/ETH01020, 2019/ETH07783). OUTLOOK is approved to have waiver of individual patient consent. TIMELY, Co-TIMELY and CONTEND participants will provide written informed consent. Final results will be disseminated through peer-reviewed journals and presented at scientific meetings.

Construction and Comprehensive Analysis of miRNAs and Target mRNAs in Longissimus dorsi Muscle of Queshan Black and Large White Pigs.

A miRNA-mRNA combination analysis was performed on the longissimus dorsi muscle of adult Queshan Black and Large White pigs by RNA-seq technology to reveal the molecular mechanism affecting pork quality traits. The sequencing results showed that 39 miRNAs were differentially expressed between Queshan Black and Large White pigs, which targeted 5234 mRNAs, and 15 differentially expressed miRNAs targeted 86 differentially expressed mRNAs. The qRT-PCR results showed that miRNAs showed similar expression patterns to RNA-seq. The GO analysis indicated that differentially expressed miRNAs with differential target mRNAs were primarily involved in biological processes such as phospholipase activity, MAP-kinase scaffold activity, lipase activity, and regulation of the extent of cell growth. The KEGG analysis also revealed that such mRNAs were significantly enriched in the ECM-receptor interaction, sphingolipid metabolism, apoptosis, PI3K-Akt signaling pathway, and AMPK signaling pathway. In addition, software predictions showed that 17 (13 of which were upregulated and four were downregulated) of 39 differentially expressed miRNAs targeted 118 negatively correlated expression mRNAs. The upregulated miRNAs contained 103 negatively correlated target mRNAs, whereas the downregulated miRNAs contained 15 negatively correlated target mRNAs. The GO analysis showed that such mRNAs were primarily involved in MAP-kinase scaffold activity, myoblast development, and peptidyl-lysine methylation, and the KEGG analysis showed significant enrichment in ECM-receptor interaction and focal adhesion. The functional enrichment analysis of miRNA target genes revealed that miR-328 was screened out as a key miRNA, and preliminary functional validation was performed. Moreover, the overexpressed miR-328 could affect the expression of proliferation-related genes, such as CDK2, CDK4, CCNB1, CCND1, CCNE1, and PCNA. These results indicated that miR-328 may regulate fat deposition and affect meat quality by influencing related pathways. This study revealed that the miRNA-mRNA regulatory axis affects fat deposition and skeletal muscle development, which provides a theoretical basis for further study on the molecular mechanism of meat quality.

CircSETBP1 Acts as a MiR-149-5p Sponge to Promote Intramuscular Fat Deposition by Regulating CRTCs.

Circular RNAs (circRNAs) appear to be crucial in the process of adipogenesis according to mounting data. CircSETBP1 is a newly discovered circRNA associated with adipogenesis. Sequencing verification and RNase R treatment have confirmed the circular nature of circSETBP1 in porcine tissue. The precise function and mechanism of circSETBP1 in adipocyte biology are still unclear. Cell counting kit-8 (CCK8), Oil red O staining, and quantitative real-time polymerase chain reaction (qRT-PCR) were employed in this investigation to reveal the functions of circSETBP1 and miR-149-5p in the growth and development of porcine intramuscular (IM) preadipocytes. CircSETBP1 overexpression accelerated cell differentiation while reducing cell proliferation. The opposite outcome was produced by overexpressing miR-149-5p. Meanwhile, circSETBP1 down-regulated the expression of miR-149-5p and miR-149-5p restrained the expression of CRTC1/CRTC2. CircSETBP1 was directly targeted by miR-149-5p, and CRTC1/CRTC2 were the target genes of miR-149-5p using bioinformatic analysis, the dual-Luciferase reporter system, and qRT-PCR. In conclusion, circSETBP1 controls the proliferation and differentiation of porcine IM preadipocytes and 3T3-L1 cells by regulating the miR-149-5p/CRTCs axis. The results of this study not only illuminate the molecular mechanism of circSETBP1/miR-149-5p involved in the deposition of porcine intramuscular fat (IMF), but they also provide a significant theoretical reference for raising quality of pork.

Identification of Estrus in Sows Based on Salivary Proteomics.

The estrus cycle of multiparous Large White sows was divided into three stages to solve the problems of heavy workload and low accuracy of the traditional estrus identification method in pig production. Saliva protein was extracted from the oral saliva of multiparous sows. Label-free quantitative proteomics was used to detect salivary proteome, and MaxQuant software was used for quality control. Results showed that 246 proteins were identified in the three stages, where 40 proteins were significantly different (p < 0.05). The total proteins identified were enriched by STEM software and the protein function was annotated by using the ClueGO plug-in in the Cytoscape software. The results were enriched to eight different trends. The annotated items were related to protein synthesis and processing and estrogen response. Gene ontology and the Kyoto Encyclopedia of Genes and Genomes enrichment analysis of differential proteins involved in the pathways and entries included oocyte meiosis, response to estradiol, and oogenesis. Further interaction analysis showed that an interaction occurred between P00355, F1SHL9, P28491, F1SDR7, F2Z558, F1RYY6, and F2Z5G3 proteins. The findings served as a basis for revealing the changes in salivary protein content in the sow estrus cycle and provided a reference for the development of an estrus identification kit/test strip in the next step.

Construction of circRNA-related ceRNA networks in longissimus dorsi muscle of Queshan Black and Large White pigs.

Circular RNA (circRNA) is a class of non-coding RNA (ncRNA) that plays an important regulatory role in various biological processes of the organisms and has a major function in muscle growth and development. However, its molecular mechanisms of how it regulates pork quality remain unclear at present. In this study, we compared the longissimus dorsi (LD) muscle expression profiles of Queshan Black (QS) and Large White (LW) pigs to explore the role of circRNAs in meat quality using transcriptome sequencing. A total of 62 differentially expressed circRNAs (DECs), including 46 up- and 16 down-regulated, 39 differentially expressed miRNAs (DEmiRNAs), including 21 up- and 18 down-regulated and 404 differentially expressed mRNAs (DEMs), including 174 up- and 230 down-regulated were identified, and most circRNAs were composed of exons. Our results indicated that the DEC parent genes and DEMs were enriched in the positive regulation of fast-twitch skeletal muscle fiber contraction, relaxation of skeletal muscle, regulation of myoblast proliferation, AMPK signaling pathway, Wnt and Jak-STAT signaling pathway. Furthermore, circSETBP1/ssc-miR-149/PIK3CD and circGUCY2C/ssc-miR-425-3p/CFL1 were selected by constructing the competitive endogenous RNA (ceRNA) regulatory network, circSETBP1, circGUCY2C, PIK3CD and CFL1 had low expression level in QS, while ssc-miR-149 and ssc-miR-425-3p had higher expression level than LW, our analysis revealed that circSETBP1, circGUCY2C, ssc-miR-149, ssc-miR-425-3p, PIK3CD and CFL1 were associated with lipid regulation, cell proliferation and differentiation, so the two ceRNAs regulatory networks may play an important role in regulating intramuscular fat (IMF) deposition, thereby affecting pork quality. In conclusion, we described the gene regulation by the circRNA-miRNA-mRNA ceRNA networks by comparing QS and LW pigs LD muscle transcriptome, and the two new circRNA-associated ceRNA regulatory networks that could help to elucidate the formation mechanism of pork quality. The results provide a theoretical basis for further understanding the genetic mechanism of meat quality formation.

MicroRNA-24 protects against myocardial ischemia-reperfusion injury via the NF-κB/TNF-α pathway.

Acute myocardial infarction (AMI) is a form of cardiomyopathy in which a blocked coronary artery leads to an irreversible loss of cardiomyocytes due to inadequate blood and oxygen supply to the distal myocardium tissues, eventually leading to heart failure. Recently, studies have revealed that microRNA (miRNA/miR)-24 has diagnostic value in the pathogenesis of AMI by affecting multiple cell processes such as cell proliferation, differentiation and apoptosis. However, the specific mechanism of miR-24 in ischemia-reperfusion injury (IRI) after AMI remains to be fully elucidated. The present study aimed to investigate the effects and mechanisms of miR-24 in IRI. In vitro, the current study detected cellular apoptosis and apoptotic-related protein expression levels in the cardiomyocyte H9C2 cell line (negative control group, model group and miRNA group) via flow cytometry and western blot analysis. In the in vivo study, rats were randomly divided into sham, model and miRNA groups. The infarct area was observed using nitro blue tetrazolium staining, pathological changes of the myocardium were detected via hematoxylin and eosin staining and TUNEL staining was used to detect cardiomyocyte apoptosis. The expression levels of related proteins were evaluated via immunohistochemistry and western blot analysis. The in vitro and in vivo results demonstrated that miR-24 significantly inhibited cardiomyocyte apoptosis compared with the model group. Concurrently, the expression levels of proteins associated with the NF-κB/TNF-α pathway (NF-κB, caspase-3, Bax, Bcl-2, TNF-α, vascular cell adhesion molecule 1, intercellular adhesion molecule 1 and monocyte chemoattractant protein-1) in the miRNA group were significantly different from the model group (P<0.001). Compared with the model group, miR-24 significantly improved pathological damage and infarct size of rat myocardium. Overall, the present results suggested that miR-24 improves myocardial injury in rats by inhibiting the NF-κB/TNF-α pathway.

Ultra-broadband polarization beam splitter with silicon subwavelength-grating waveguides.

An ultra-broadband polarization beam splitter (PBS) with low excess loss (EL) and a high extinction ratio (ER) is proposed and demonstrated for the case with 340 nm thick silicon-on-insulator waveguides. Here the PBS is realized by using cascaded adiabatic dual-core tapers, which consist of a strip core and a subwavelength-grating core. For the designed PBS, which has a 33.6 µm long mode-evolution region, the ELs are ${ \lt }{0.3}\;{\rm dB}$<0.3dB, and the ERs are ${ \gt }{20}\;{\rm dB}$>20dB for both TE and TM polarizations in an ultra-broad bandwidth of ${ \gt }{270}\;{\rm nm}$>270nm (1400-1670 nm) in theory. For the fabricated PBS, the measured bandwidths for achieving ERs of ${\sim}{20}$∼20 and ${\sim}{25}\;{\rm dB}$∼25dB are 240 and 220 nm, while the 1 dB bandwidth is as large as 230 nm, which are the largest bandwidths reported to date, to the best of our knowledge.

Silicon-based hybrid demultiplexer for wavelength- and mode-division multiplexing.

A silicon-based hybrid demultiplexer for wavelength-division multiplexing (WDM) and mode-division multiplexing (MDM) is proposed and demonstrated by integrating an M-channel-mode demultiplexer and N-channel WDM filters based on microring resonators (MRRs) with box-like responses. For the mode demultiplexer, the 2k-th output port is connected with the (2k+1)-th output port through the bus waveguide for the k-th MRR array, so that each MRR-based optical filter works bi-directionally and provides two drop ports. As an example, a 32-channel hybrid MDM-WDM demultiplexer is realized by integrating a 4-channel mode demultiplexer based on dual-core adiabatic tapers and two bi-directional MRR-based WDM filters with eight wavelength-channels. For the fabricated hybrid demultiplexer, the excess loss is 0.5-5 dB, the intermode cross talk is -16.5 to -23.5 dB, and the cross talks between the adjacent and nonadjacent wavelength channels are about -25 dB and -35 dB, respectively.

Biodegradation of Feather Waste Keratin by the Keratin-Degrading Strain Bacillus subtilis 8.

Bacillus subtilis 8 is highly efficient at degrading feather keratin. We observed integrated feather degradation over the course of 48 h in basic culture medium while studying the entire process with scanning electron microscopy. Large amounts of ammonia, sulfite, and L-cysteic acid were detected in the fermented liquid. In addition, four enzymes (gamma-glutamyltranspeptidase, peptidase T, serine protease, and cystathionine gamma-synthase) were identified that play an important role in this degradation pathway, all of which were verified with molecular cloning and prokaryotic expression. To the best of our knowledge, this report is the first to demonstrate that cystathionine gamma-synthase secreted by B. subtilis 8 is involved in the decomposition of feather keratin. This study provides new data characterizing the molecular mechanism of feather degradation by bacteria, as well as potential guidance for future industrial utilization of waste keratin.

Improved thermostability and enzyme activity of a recombinant phyA mutant phytase from Aspergillus niger N25 by directed evolution and site-directed mutagenesis.

We previously constructed three recombinant phyA mutant strains (PP-NPm-8, PP-NPep-6A and I44E/T252R-PhyA), showing improved catalytic efficiency or thermostability of Aspergillus niger N25 phytase, by error-prone PCR or site-directed mutagenesis. In this study, directed evolution and site-directed mutagenesis were further applied to improve the modified phytase properties. After one-round error-prone PCR for phytase gene of PP-NPep-6A, a single transformant, T195L/Q368E/F376Y, was obtained with the significant improvements in catalytic efficiency and thermostability. The phytase gene of T195L/Q368E/F376Y, combined with the previous mutant phytase genes of PP-NPep-6A, PP-NPm-8 and I44E/T252R-PhyA, was then sequentially modified by DNA shuffling. Three genetically engineered strains with desirable properties were then obtained, namedQ172R, Q172R/K432R andQ368E/K432R. Among them, Q172R/K432R showed the highest thermostability with the longest half-life and the greatest remaining phytase activity after heat treatment, while Q368E/K432R showed the highest catalytic activity. Five substitutions (Q172R, T195L, Q368E, F376Y, K432R) identified from random mutagenesis were added sequentially to the phytase gene of PP-NPep-6A to investigate how the mutant sites influence the properties of phytase. Characterization and structural analysis demonstrated that these mutations could produce cumulative or synergistic improvements in thermostability or catalytic efficiency of phytase.

Compact polarization beam splitter for silicon photonic integrated circuits with a 340-nm-thick silicon core layer.

A polarization beam splitter (PBS) is proposed and realized for silicon photonic integrated circuits with a 340-nm-thick silicon core layer by introducing an asymmetric directional coupler (ADC), which consists of a silicon-on-insulator (SOI) nanowire and a subwavelength grating (SWG) waveguide. The SWG is introduced to provide an optical waveguide which has much higher birefringence than a regular 340-nm-thick SOI nanowire, so that it is possible to make the phase-matching condition satisfied for TE polarization only in the present design when the waveguide dimensions are optimized. Meanwhile, there is a significant phase mismatching for TM polarization automatically. In this way, the present ADC enables strong polarization selectivity to realize a PBS that separates TE and TM polarizations to the cross and through ports, respectively. The realized PBS has a length of ∼2 µm for the coupling region. For the fabricated PBS, the extinction ratio (ER) is 15-30 dB and the excess loss is 0.2-2.6 dB for TE polarization while the ER is 20-27 dB and the excess loss is 0.3-2.8 dB for TM polarization when operating in the wavelength range of 1520-1580 nm.