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Vertical cavity surface emitting lasers (VCSELs) have emerged as a versatile and promising platform for developing advanced integrated photonic devices and systems due to their low power consumption, high modulation bandwidth, small footprint, excellent scalability, and compatibility with monolithic integration. By combining these unique capabilities of VCSELs with the functionalities offered by micro/nano optical structures (e.g. metasurfaces), it enables various versatile energy-efficient integrated photonic devices and systems with compact size, enhanced performance, and improved reliability and functionality. This review provides a comprehensive overview of the state-of-the-art versatile integrated photonic devices/systems based on VCSELs, including photonic neural networks, vortex beam emitters, holographic devices, beam deflectors, atomic sensors, and biosensors. By leveraging the capabilities of VCSELs, these integrated photonic devices/systems open up new opportunities in various fields, including artificial intelligence, large-capacity optical communication, imaging, biosensing, and so on. Through this comprehensive review, we aim to provide a detailed understanding of the pivotal role played by VCSELs in integrated photonics and highlight their significance in advancing the field towards efficient, compact, and versatile photonic solutions., (© 2024. The Author(s).)

Holography technology is considered the ultimate three-dimensional (3D) visualization technology in the future. However, conventional methods for achieving holography generally utilize discrete optical components and off-chip laser sources, resulting in a large size and high complexity, which are undesirable for practical applications. In this Letter, chip-scale integrated holographic devices are realized by integrating top-emitting vertical cavity surface emitting lasers (VCSELs) with micro holograms printed by 3D femtosecond laser nanoprinting technology. The VCSELs are designed to operate in a single fundamental mode with a Gaussian emission profile. Then the Gaussian beams are phase-modulated by the integrated micro holograms designed by the Gerchberg-Saxton (GS) algorithm and the target holographic images can be displayed behind the holograms. Such integrated holographic devices are of micron size and can be easily scaled into arrays with arbitrary channels on-demand, which are important for achieving miniaturized and portable holographic imaging systems.

Enterovirus 71 (EV71) is one of the major pathogens causing hand, foot, and mouth disease in children under 5 years old, which can result in severe neurological complications and even death. Due to limited treatments for EV71 infection, the identification of novel host factors and elucidation of mechanisms involved will help to counter this viral infection. N-terminal acetyltransferase 6 (NAT6) was identified as an essential host factor for EV71 infection with genome-wide CRISPR/Cas9 screening. NAT6 facilitates EV71 viral replication depending on its acetyltransferase activity but has little effect on viral release. In addition, NAT6 is also required for Echovirus 7 and coxsackievirus B5 infection, suggesting it might be a pan-enterovirus host factor. We further demonstrated that NAT6 is required for Golgi integrity and viral replication organelle (RO) biogenesis. NAT6 knockout significantly inhibited phosphatidylinositol 4-kinase IIIβ (PI4KB) expression and PI4P production, both of which are key host factors for enterovirus infection and RO biogenesis. Further mechanism studies confirmed that NAT6 formed a complex with its substrate actin and one of the PI4KB recruiters-acyl-coenzyme A binding domain containing 3 (ACBD3). Through modulating actin dynamics, NAT6 maintained the integrity of the Golgi and the stability of ACBD3, thereby enhancing EV71 infection. Collectively, these results uncovered a novel mechanism of N-acetyltransferase supporting EV71 infection.IMPORTANCEEnterovirus 71 (EV71) is an important pathogen for children under the age of five, and currently, no effective treatment is available. Elucidating the mechanism of novel host factors supporting viral infection will reveal potential antiviral targets and aid antiviral development. Here, we demonstrated that a novel N-acetyltransferase, NAT6, is an essential host factor for EV71 replication. NAT6 could promote viral replication organelle (RO) formation to enhance viral replication. The formation of enterovirus ROs requires numerous host factors, including acyl-coenzyme A binding domain containing 3 (ACBD3) and phosphatidylinositol 4-kinase IIIβ (PI4KB). NAT6 could stabilize the PI4KB recruiter, ACBD3, by inhibiting the autophagy degradation pathway. This study provides a fresh insight into the relationship between N-acetyltransferase and viral infection., Competing Interests: The authors declare no conflict of interest.

Given the serious neurological complications and deaths associated with enterovirus 71 (EV71) infection, there is an urgent need to develop effective antivirals against this viral infection. In this study, we demonstrated that two Cathelicidin-derived peptides, LL-18 and FF-18 were more potent against EV71 infection than the parent peptide LL-37, which is the mature and processed form of Cathelicidin. These peptides could directly bind to the EV71 virus particles, but not to coxsackievirus, indicative of their high specificity. The binding of peptides with the virus surface occupied the viral canyon region in a way that could block virus-receptor interactions and inhibit viral uncoating. In addition, these peptide analogues could also relieve the deleterious effect of EV71 infection in vivo. Therefore, Cathelicidin-derived peptides might be excellent candidates for further development of antivirals to treat EV71 infection., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Fan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Reductions in Na+-K+-ATPase (NKA) activity and expression are often observed in the progress of various reason-induced heart failure (HF). However, NKA α1 mutation or knockdown cannot cause spontaneous heart disease. Whether the abnormal NKA α1 directly contributes to HF pathogenesis remains unknown. Here, we challenge NKA α1+/- mice with isoproterenol to evaluate the role of NKA α1 haploinsufficiency in isoproterenol (ISO)-induced cardiac dysfunction. Genetic knockdown of NKA α1 accelerated ISO-induced cardiac cell hypertrophy, heart fibrosis, and dysfunction. Further studies revealed decreased Krebs cycle, fatty acid oxidation, and mitochondrial OXPHOS in the hearts of NKA α1+/- mice challenged with ISO. In ISO-treated conditions, inhibition of NKA elevated cytosolic Na+, further reduced mitochondrial Ca2+ via mNCE, and then finally down-regulated cardiac cell energy metabolism. In addition, a supplement of DRm217 alleviated ISO-induced heart dysfunction, mitigated cardiac remodeling, and improved cytosolic Na+ and Ca2+ elevation and mitochondrial Ca2+ depression in the NKA α1+/- mouse model. The findings suggest that targeting NKA and mitochondria Ca2+ could be a promising strategy in the treatment of heart disease., (© 2024 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Aspartic proteases (ASPs) are important hydrolases for parasitic invasion of host tissues or cells. This was the first study on Demodex ASP. First, the complete coding sequence (CDS) was amplified, cloned and sequenced. Then, the protein physical and chemical properties was analysed. Finally, the recombinant plasmid, expression and purification system was established. Results showed that the lengths of CDS of Demodex folliculorum and D. brevis were 1161 and 1173 bp, respectively. The molecular weight of the protein was approximately 40 KDa. It contained an aspartic acid residue, a substrate-binding site and signal peptide, yet lacked a transmembrane domain and was located in the membrane or extracellular matrix. The phylogenetic and conserved motif analyses showed that D. folliculorum and D. brevis clustered separately and then formed a single branch, which finally clustered with other Acariformes species. The prokaryotic expression systems for recombinant ASP with His-tag (rASP-His) and GST-tag (rASP-GST) were constructed. The inclusion bodies of rASP-His were renaturated by gradient urea and purified using NI beads, while those of rASP-GST were renaturated by sarkosyl and Triton X-100 and purified using GST beads. Conclusively, the prokaryotic expression and purification system of Demodex rASP was successfully established for further pathogenic mechanism research., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Vertical-cavity surface-emitting lasers (VCSELs) represent an attractive light source to integrate with OAM structures to realize chip-scale vortex lasers. Although pioneering endeavors of VCSEL-based vortex lasers have been reported, they cannot achieve large topological charges (less than l = 5) due to the insufficient space-bandwidth product (SBP) caused by the inherent limited device size. Here, by integrating a nanoprinted OAM phase structure on the VCSELs, we demonstrate a vortex microlaser with a low threshold and simple structure. A monolithic microlaser array with addressable control of vortex beams with different topological charges ( l = 1 to l = 5) was achieved. Nanoprinting offers high degrees of freedom for the manipulation of spatial structures. To address the challenge of insufficient SBP, two-layer cascaded spiral phase plates were designed. Thereby, a vortex beam with l = 15 and mode purity of 83.7% was obtained. Our work paves the way for future chip-scale OAM-based information multiplexing with more channels.

The human astrovirus (HAstV) is a non-enveloped, single-stranded RNA virus that is a common cause of gastroenteritis. Most non-enveloped viruses use membrane disruption to deliver the viral genome into a host cell after virus uptake. The virus-host factors that allow for HAstV cell entry are currently unknown but thought to be associated with the host-protease-mediated viral maturation. Using in vitro liposome disruption analysis, we identified a trypsin-dependent lipid disruption activity in the capsid protein of HAstV serotype 8. This function was further localized to the P1 domain of the viral capsid core, which was both necessary and sufficient for membrane disruption. Site-directed mutagenesis identified a cluster of four trypsin cleavage sites necessary to retain the lipid disruption activity, which is likely attributed to a short stretch of sequence ending at arginine 313 based on mass spectrometry of liposome-associated peptides. The membrane disruption activity was conserved across several other HAstVs, including the emerging VA2 strain, and effective against a wide range of lipid identities. This work provides key functional insight into the protease maturation process essential to HAstV infectivity and presents a method to investigate membrane penetration by non-enveloped viruses in vitro . IMPORTANCE Human astroviruses (HAstVs) are an understudied family of viruses that cause mild gastroenteritis but have recent cases associated with a more severe neural pathogenesis. Many important elements of the HAstV life cycle are not well understood, and further elucidating them can help understand the various forms of HAstV pathogenesis. In this study, we utilized an in vitro liposome-based assay to describe and characterize a previously unreported lipid disruption activity. This activity is dependent on the protease cleavage of key sites in HAstV capsid core and can be controlled by site-directed mutagenesis. Our group observed this activity in multiple strains of HAstV and in multiple lipid conditions, indicating this may be a conserved activity across the AstV family. The discovery of this function provides insight into HAstV cellular entry, pathogenesis, and a possible target for future therapeutics., Competing Interests: The authors declare no conflict of interest.

Coxsackievirus belongs to the Picornaviridae family and is one of the major pathogens that cause hand, foot and mouth disease (HFMD) in infants and children with potential serious complications and even deaths. The pathogenesis of this virus is not fully elucidated and no vaccine or antiviral drug has been approved. In this study, a full-length infectious cDNA clone of coxsackievirus B5 virus was assembled and the recombinant virus displayed similar growth kinetics and ability to cause cytopathic effects as the parental virus. Luciferase reporter was then incorporated to generate both full-length and subgenomic replicon (SGR) reporter viruses. The full-length reporter virus is suitable for high-throughput antiviral screening, while the SGR is a useful tool to study viral-host interactions. More importantly, the full-length reporter virus has also been shown to infect the suckling mouse model and the reporter gene could be detected using an in vivo imaging system, thus providing a powerful tool to track viruses in vivo. In summary, we have generated coxsackievirus B5 reporter viruses and provided unique tools for studying virus-host interactions in vitro and in vivo as well as for high-throughput screenings (HTS) to identify novel antivirals., (Copyright © 2023 The Authors. Publishing services by Elsevier B.V. All rights reserved.)

Acinetobacter baumannii is a critical biofilm-forming pathogen that has presented great challenges in the clinic due to multidrug resistance. Thus, new methods of intervention are needed to control biofilm-associated infections. In this study, among three tested Lactobacillus species, Lactobacillus rhamnosus showed significant antimaturation and antiadherence effects against A. baumannii biofilm. Lactic acid (LA) and acetic acid (AA) were the most effective antibiofilm biosurfactants (BSs) produced by L. rhamnosus. This antibiofilm phenomenon produced by LA and AA was due to the strong bactericidal effect, which worked from very early time points, as determined by colony enumeration and confocal laser scanning microscope. The cell destruction of A. baumannii appeared in both the cell envelope and cytoplasm. A discontinuous cell envelope, the leakage of cell contents, and the increased extracellular activity of ATPase demonstrated the disruption of the cell membrane by LA and AA. These effects also demonstrated the occurrence of protein lysis. In addition, bacterial DNA interacted with and was damaged by LA and AA, resulting in significantly reduced expression of biofilm and DNA repair genes. The results highlight the possibility and importance of using probiotics in clinical prevention. Probiotics can be utilized as novel biocides to block and decrease biofilm formation and microbial contamination in medical equipment and during the treatment of infections. IMPORTANCE A. baumannii biofilm is a significant virulence factor that causes the biofilm colonization of invasive illnesses. Rising bacterial resistance to synthetic antimicrobials has prompted researchers to look at natural alternatives, such as probiotics and their derivatives. In this study, L. rhamnosus and its BSs (LA and AA) demonstrated remarkable antibiofilm and antimicrobial characteristics, with a significant inhibitory effect on A. baumannii. These effects were achieved by several mechanisms, including the disruption of the cell envelope membrane, protein lysis, reduced expression of biofilm-related genes, and destruction of bacterial DNA. The results provide support for the possibility of using probiotics and their derivatives in the clinical prevention and therapy of A. baumannii infections.

Reduced Na + /K + -ATPase (NKA) activity and NKAα1 expression are engaged in the pathologies of renal diseases. NKA-mediated Src activation is not the only reason for NKA-related renal fibrosis. In this study, we found that genetic reduction of NKAα1 exhibited exacerbated tubulointerstitial lesions and fibrosis in the UUO mice model. Activation of NKAα1 with an antibody against the extracellular DR region of the NKAα1 subunit (DRm217) prevented UUO-induced tubulointerstitial lesions, preserved kidney function, and decrease renal fibrosis. Further studies revealed that NKAα1 deficiency mice exhibited high inflammation factors expression when they suffered UUO surgery, compared with NKAα1 +/+ (WT) mice. DRm217 alleviated inflammatory cell infiltration, suppress NF-κB phosphorylation, and decreased inflammatory factors expression in the UUO mice model. Released HMGB1 can trigger the inflammatory response and contribute to renal fibrosis. Knockdown of NKA in renal tubular cells or in NKAα1 +/- mice was associated with more susceptibility to HMGB1 release in the UUO mice model. DRm217 exerted its antifibrotic effect via inhibiting HMGB1 release. Furthermore, AMPK activation participates in the effect of DRm217 on inhibiting HMGB1 release. Our findings suggest that NKAα1 is a regulator of renal fibrosis and its DR-region is a novel target on it., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

As the greatest defense organ of the body, the skin is exposed to endogenous and external stressors that produce reactive oxygen species (ROS). When the antioxidant system of the body fails to eliminate ROS, oxidative stress is initiated, which results in skin cellular senescence, inflammation, and cancer. Two main possible mechanisms underlie oxidative stress-induced skin cellular senescence, inflammation, and cancer. One mechanism is that ROS directly degrade biological macromolecules, including proteins, DNA, and lipids, that are essential for cell metabolism, survival, and genetics. Another one is that ROS mediate signaling pathways, such as MAPK, JAK/STAT, PI3K/AKT/mTOR, NF-κB, Nrf2, and SIRT1/FOXO, affecting cytokine release and enzyme expression. As natural antioxidants, plant polyphenols are safe and exhibit a therapeutic potential. We here discuss in detail the therapeutic potential of selected polyphenolic compounds and outline relevant molecular targets. Polyphenols selected here for study according to their structural classification include curcumin, catechins, resveratrol, quercetin, ellagic acid, and procyanidins. Finally, the latest delivery of plant polyphenols to the skin (taking curcumin as an example) and the current status of clinical research are summarized, providing a theoretical foundation for future clinical research and the generation of new pharmaceuticals and cosmetics.

This study aimed to develop an effective method for the expression and purification of the Dermatophagoides farinae serpin protein and to establish an experimental foundation for elucidating its role in the temperature stress response. The total RNA of D. farinae was extracted, and specific primers were designed for serpin amplification. Serpin was joined with pET32a vector and transformed into BL21 (DE3) cells. Expression of recombinant proteins was induced. Proteins were extracted by enzymatic lysis or enzymatic lysis combined with ultrasonication. Recombinant proteins were purified by Ni-NTA method. SDS-PAGE was conducted to evaluate protein expression, extraction, and purification efficiency. Agarose gel electrophoresis and sequencing analysis showed that the amplified serpin open reading frame was 1284 bp, encoding a hydrophilic and stable protein with a relative molecular weight of 48.30 kD. SDS-PAGE demonstrated that there was a specific band at 55-70 kD, which was consistent with the predicted size of the recombinant pET32a-Serpin protein. Enzymatic lysis combined with 30% ultrasonic power promoted the release of soluble protein more effectively than enzymatic lysis alone. 16 °C for 4 h was optimal for inducing expression. The optimal imidazole concentrations for washing non-His-tagged protein and eluting His-tagged protein were determined to be 20 mM and 200 mM, respectively. In this study, A prokaryotic expression and purification system for the D. farinae serpin protein was successfully established, providing a technical reference for functional gene research in mites at the protein level., (Copyright © 2022 Elsevier Inc. All rights reserved.)

The recent COVID-19 pandemic poses a global health emergency. Cellular entry of the causative agent SARS-CoV-2 is mediated by its spike protein interacting with cellular receptor-human angiotensin converting enzyme 2 (ACE2). Here, by using lentivirus based pseudotypes bearing spike protein, we demonstrated that entry of SARS-CoV-2 into host cells was dependent on clathrin-mediated endocytosis, and phosphoinositides played essential roles during this process. In addition, we showed that the intracellular domain and the catalytic activity of ACE2 were not required for efficient virus entry. Finally, we showed that the current predominant Delta variant, although with high infectivity and high syncytium formation, also entered cells through clathrin-mediated endocytosis. These results provide new insights into SARS-CoV-2 cellular entry and present proof of principle that targeting viral entry could be an effective way to treat different variant infections., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest., (Copyright © 2022 The Authors. Publishing services by Elsevier B.V. All rights reserved.)

Enterovirus 71 (EV71) is deemed a reemergent pathogen, with recent outbreaks worldwide. EV71 infection causes hand, foot, and mouth disease (HFMD) and has been associated with severe cardiac and central nervous system complications and even death. Viruses need host factors to complete their life cycle; therefore, the identification of the host factors for EV71 infection is pivotal to new antiviral research. Emerging evidence has highlighted the importance of protein acetylation during infection by various human viruses. The endoplasmic reticulum (ER), as the prominent organelle of EV71 replication, also has a unique acetylation regulation mechanism. However, the pathogenesis of EV71 and its relationship with the ER-based acetylation machinery are not fully understood. In this study, we demonstrated for the first time that the ER-resident acetyltransferase N -acetyltransferase 8 (NAT8) is a host factor for EV71 infection. Inhibiting NAT8 with CRISPR or a small compound significantly suppressed EV71 infection in SK-N-SH cells. NAT8 promoted EV71 replication in an acetyltransferase-activity-dependent manner. Additionally, we found that NAT8 facilitates EV71 infection by interacting with EV71 2B, 3AB, and 3C proteins and increasing the stability of these proteins. These results uncovered a novel function of NAT8 and elucidated a new mechanism underlying the regulation of EV71 replication. IMPORTANCE EV71 is one of the most common pathogens causing HFMD in young children, and some patients experience severe or fatal neurological consequences. To ensure efficient replication, the virus must hijack multiple host factors for its own benefit. Here, we show that the ER-resident acetyltransferase NAT8 is a host factor for EV71 infection. EV71 fails to complete its infection in various cells in the absence of NAT8. We further show that NAT8 benefits EV71 replication in an acetyltransferase-activity-dependent manner. Finally, we show that NAT8 facilitates EV71 infection by interacting with EV71 2B, 3AB, and 3C proteins and increasing the stability of these proteins. These results uncovered a novel function of NAT8 in EV71 infection and elucidated a new mechanism underlying the regulation of EV71 replication.

GeSn materials have attracted considerable attention for their tunable band structures and high carrier mobilities, which serve well for future photonic and electronic applications. This research presents a novel method to incorporate Sn content as high as 18% into GeSn layers grown at 285-320 °C by using SnCl 4 and GeH 4 precursors. A series of characterizations were performed to study the material quality, strain, surface roughness, and optical properties of GeSn layers. The Sn content could be calculated using lattice mismatch parameters provided by X-ray analysis. The strain in GeSn layers was modulated from fully strained to partially strained by etching Ge buffer into Ge/GeSn heterostructures . In this study, two categories of samples were prepared when the Ge buffer was either laterally etched onto Si wafers, or vertically etched Ge/GeSnOI wafers which bonded to the oxide. In the latter case, the Ge buffer was initially etched step-by-step for the strain relaxation study. Meanwhile, the Ge/GeSn heterostructure in the first group of samples was patterned into the form of micro-disks. The Ge buffer was selectively etched by using a CF 4 /O 2 gas mixture using a plasma etch tool. Fully or partially relaxed GeSn micro-disks showed photoluminescence (PL) at room temperature. PL results showed that red-shift was clearly observed from the GeSn micro-disk structure, indicating that the compressive strain in the as-grown GeSn material was partially released. Our results pave the path for the growth of high quality GeSn layers with high Sn content, in addition to methods for modulating the strain for lasing and detection of short-wavelength infrared at room temperature.

In this study, ultrasonic-assisted cellulase extraction (UCE) was applied to extract flavonoids and polyphenols from the Nymphaea hybrid flower. The extraction conditions were optimized using the response surface method (RSM) coupled with a Box-Behnken design. The crude extract of Nymphaea hybrid (NHE) was further purified using AB-8 macroporous resins, and the purified extract (NHEP) was characterized by FTIR and HPLC. In vitro activity determination by chemical method showed that NHEP displayed strong free radical scavenging abilities against the DPPH and ABTS radicals, good reduction power, and hyaluronidase inhibition. The cell viability by CCK-8 assays showed that NHEP had no significant cytotoxicity for B16 and HaCaT cells when the concentration was below 100 μg/mL and 120 μg/mL, respectively. NHEP with a concentration of 20-160 μg/mL can more effectively reduce the ROS level in H 2 O 2 damaged HaCaT cells compared with 10 μg/mL of VC. The 40 μg/mL of NHEP had similar activity against intracellular melanin production in the B16 melanoma cells compared with 20 μg/mL Kojic acid. Good activities of antioxidation, whitening and protective effect against H 2 O 2 -induced oxidative damage promote the potential for NHEP as a functional raw material in the field of cosmetics and medicine.

Acinetobacter baumannii is an important pathogen in clinical. The factors of biofilm formation, antibiotic resistance and motility contribute great to A. baumannii in persisting in stressed environment, and further leads to nosocomial infections. 70 A. baumannii clinical isolates were investigated for their clinical characteristics of infection. Among the tested strains, 54 (77.1%) isolates were obtained from ICUs, with the frequency of multidrug-resistance (MDR) at 55.7%, and that of extensively drug-resistance (XDR) at 31.4%. 97.1% of the clinical isolates could form biofilms, in which 4.3% possessed weak biofilm formation ability, while 41.4% and 51.4% were moderate and strong biofilm producers, respectively. A strong correlation between antibiotic resistance and biofilm formation ability was found that all the resistant strains could form biofilms, with the majority in moderate and strong levels, but 2.9% sensitive isolates had no such ability. However, the sensitive strains that could produce biofilms showed stronger biofilm formation capacity in the early stage before 24 h compared to the resistant isolates, though they became weaker afterwards. 24 biofilm-related genes and two bla OXA genes were found in both biofilm-forming and non-biofilm-forming strains, but with higher prevalence in the strains that could produce biofilms. No correlation was detected between twitching motility with antibiotic susceptibility or biofilm formation. These results raised a viewpoint that examining timepoint is a key factor for determining the biofilm formation ability, and further highlighted the importance of the appropriate surveillance and control measures in preventing the emergence and transmission of MDR and XDR A. baumannii., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Enterovirus 71 (EV71) is a neurotropic pathogen that causes hand, foot, and mouth disease (HFMD) and it has been consistently associated with severe neurological, cardiac, and respiratory complications. Yet there is no specific treatment for this virus and we still know little about the viral pathogenesis. In this study, we first generated an infectious cDNA clone of EV71 virus from a patient virus strain and made a full-length virus with a NanoLuc reporter gene through reverse genetic approaches. The reporter gene of this virus is genetically stable when passaging in cells and could be used for antiviral testing. In addition, we also made subgenomic replicons (SGRs) of EV71, which lacks part of the structural genes dispensable for viral replication and showed that SGR can be used for viral replication study. Overall, these reporter viral systems are useful tools for EV71 pathogenesis study and antiviral screening., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Yang, Zhao, Xun, Ma and Wang.)

We apply the antenna coherence theory in order to evaluate characteristic behavior of phase-coherent VCSEL arrays. Large 19-element phase-locked VCSEL arrays with a near-diffraction-limited beam were firstly realized using proton implantation technology. The central lobe intensity is about four times that of side lobes in far-field patterns. The angular full width at half maximum (FWHM) of the far field lobes is only 1.42 degrees. A good matching between theory and experiment opens new perspectives for optimizing devices.

Antimicrobial peptides (AMPs) exhibit multiple activities against bacteria and fungi. A bovine myeloid antimicrobial peptide (BMAP-28) belongs to the cathelicidin-derived AMPs and has antimicrobial activity. Due to the rapidly increasing number of infections and outbreaks caused by pan-drug-resistant Acinetobacter baumannii (PDRAB), we sought to determine whether BMAP-28 and its 4 analog peptides (A837, A838, A839, and A840) have antimicrobial activity against PDRAB. Furthermore, we clarified the possible mechanism of inhibition by which of BMAP-28 acts against PDRAB. In the current study, we examined the inhibitory effect of BMAP-28 and its 4 analog peptides on the growth of PDRAB through minimal inhibitory concentration (MIC) analysis and short time killing assays. We also evaluated the effects of BMAP-28 and its analogs on the bacterial cell surface through the use of field emission scanning electron microscopy (FE-SEM). In order to determine the inhibitory mechanism of BMAP-28, we examined the interaction between BMAP-28 and outer membrane proteins (OMPs), especially the interaction between BMAP-28 and A. baumannii OmpA (AbOmpA), which is the main component of OMPs, by using a quartz crystal microbalance (QCM). BMAP-28 and its 4 analogs were effective in inhibiting the growth of PDRAB and had rapid killing ability. BMAP-28 showed exceptionally strong and rapid inhibitory effects on PDRAB when compared to the other peptides and was also shown to cause damage to the cell surface of PDRAB. Moreover, QCM analysis provided evidence of potential interaction between BMAP-28 and AbOmpA. These data indicate that BMAP-28 is a promising candidate for the treatment of PDRAB infections and that its inhibitory effects were related with its binding to AbOmpA.

Na + -K + -ATPase has close relationship with myocardial ischemia/reperfusion (IR) injury. Activation of Na + -K + -ATPase with its DR region specific antibody produces cardioprotective effect. In this study, we aimed to explore whether DRm217, a proved DR region specific antibody, could protect myocardial cells against IR injury and uncover the mechanisms under it. By employing H9c2 myocardial cell and SD rat, we found that DRm217 protected cardiac cells against IR-induced cell injury and apoptosis. DRm217 produced protective effect via stabilizing Na + -K + -ATPase membrane expression and inhibiting Na + -K + -ATPase/Src/NADPH oxidase dependent ROS accumulation. PI3K/Akt and ERK1/2 participated in DRm217-induced cardiomyocyte survival, but not in DRm217-related ROS reduction. Therefore, DRm217 can be used as a potential cardioprotective adjuvant in myocardial IR therapy and interference of Na + -K + -ATPase/ROS pathway will be a promising modality for clinical myocardial IR therapy., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Objective: To construct an expression vector of anti-MM scFv-tP fusion protein and test its expression efficiency and function., Methods: The truncated protamine (tP) gene sequence was added to the gene of single chain antibody against the specific antigen on the surface of malignant melanoma tumor cells using PCR. A GST-fusion expression vector was constructed and the soluable protein was expressed in the E.coli system. After cleavage and purification, the purified fusion protein was obtained. The binding activity of Anti-MM scFv-tP and siRNA was detected by EMSA. Flow cytometry and confocal microscopy were used to detect the cell surface antigen binding activity of the fusion protein., Results: The expression vector of Anti-MM scFv-tP fusion protein was successfully constructed. The soluable protein could be expressed in the E.coli system, and the purified fusion protein was obtained. The anti-MM scFv-tP fusion protein retained siRNA binding ability and could directly target malignant melanoma (MM) LiBr cells., Conclusion: The recombinant GST- Anti-MM-scFv-tp expression vector was successfully constructed. The fusion protein retains siRNA binding ability and can directly target LiBr cells to provide a reliable tool for further study.

Depressed Na + /K + -ATPase activity has long been reported to be involved in diabetic-related cardiomyocyte death and cardiac dysfunction. However, the nature of directly regulating Na + -K + -ATPase in diabetic-related myocardial diseases remains unknown. Hyperglycemia is believed as one of major factors responsible for diabetic-related myocardial apoptosis and dysfunction. In this study, whether inhibiting Na + -K + -ATPase by ouabain or activating Na + -K + -ATPase by DRm217 has functions on high glucose (HG) -induced myocardial injury was investigated. Here we found that addition of DRm217 or ouabain to HG-treated cells had opposite effects. DRm217 decreased but ouabain increased HG-induced cell injury and apoptosis. This was mediated by changing Na + -K + -ATPase activity and Na + -K + -ATPase cell surface expression. The inhibition of Na + -K + -ATPase endocytosis alleviated HG-induced ROS accumulation. Na + -K + -ATPase·c-Src dependent NADPH oxidase/ROS pathway was also involved in the effects of ouabain and DRm217 on HG-induced cell injury. These novel results may help us to understand the important role of the Na + -K + -ATPase in diabetic cardiovascular diseases., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Although gene therapy has brought new insights into the treatment of malignant melanoma, targeting delivery of nucleic acid which targets critical oncogene/anti-oncogene in vivo is still a bottleneck in the therapeutic application. Our previous in vitro studies have found that the oncogene Livin could serve as a potential molecular target by small interfering RNA for gene therapy of malignant melanoma. However, how to transport Livin small interfering RNA into malignant melanoma cells specifically and efficiently in vivo needs further investigation. Cumulative evidence has suggested that single-chain antibody-mediated small interfering RNA targeted delivery is an effective way to silence specific genes in human cancer cells. Indeed, this study designed a protamine-single-chain antibody fusion protein, anti-MM scFv-tP, to deliver Livin small interfering RNA into LiBr cells. Further experiments confirmed the induction of cell apoptosis and suppression of cell proliferation by anti-MM scFv-tP in LiBr cells, along with efficient silence of Livin gene both in vitro and in vivo. Altogether, our findings provide a feasible approach to transport Livin small interfering RNA to malignant melanoma cells which would be a new therapeutic strategy for combating malignant melanoma.

Reduced Na + -K + -ATPase activity has close relationship with cardiomyocyte death. Reactive oxygen species (ROS) also plays an important role in cardiac cell damage. It has been proved that Na + -K + -ATPase and ROS form a feed-forward amplifier. The aim of this study was to explore whether DRm217, a proved Na + /K + -ATPase's DR-region specific monoclonal antibody and direct activator, could disrupt Na + -K + -ATPase/ROS amplifier and protect cardiac cells from ROS-induced injury. We found that DRm217 protected myocardial cells against hydrogen peroxide (H 2 O 2 )-induced cardiac cell injury and mitochondrial dysfunction. DRm217 also alleviated the effect of H 2 O 2 on inhibition of Na + -K + -ATPase activity, Na + -K + -ATPase cell surface expression, and Src phosphorylation. H 2 O 2 -treatment increased intracellular ROS, mitochondrial ROS and induced intracellular Ca 2+ , mitochondrial Ca 2+ overload. DRm217 closed Na + -K + -ATPase/ROS amplifier, alleviated Ca 2+ accumulation and finally inhibited ROS and mitochondrial ROS generation. These novel results may help us to understand the important role of the Na + -K + -ATPase in oxidative stress and oxidative stress-related disease.

Hyperglycemia is one of the major factors responsible for the myocardial apoptosis and dysfunction in diabetes. Many studies have proved that there is a close relationship between decreased Na + /K + -ATPase activity and diabetic cardiomyopathy. However, the effect of directly activated Na + /K + -ATPase on high glucose-induced myocardial injury is still unknown. Here we found that DRm217, a Na + /K + -ATPase's DR-region specific monoclonal antibody and direct activator, could prevent high glucose-induced H9c2 cell injury, reactive oxygen species (ROS) release, and mitochondrial dysfunction. High glucose-treatment decreased Na + /K + -ATPase activity and increased intracellular Ca 2+ level, whereas DRm217 increased Na + /K + -ATPase activity and alleviated Ca 2+ overload. Inhibition of Ca 2+ overload or closing sodium calcium exchanger (NCX channel) could reverse high glucose-induced ROS increasing and cell injury. In addition, DRm217 could significantly attenuate high glucose-induced p38, JNK and ERK1/2 phosphorylation, which were involved in high glucose-induced cell injury and ROS accumulation. Our findings suggest that DRm217 may protect against the deleterious effects of high glucose in the heart. Prevention of high glucose-induced myocardial cell injury by specific Na + /K + -ATPase activator may be an attractive therapeutic option., (© The Author 2016. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Implant-defined vertical-cavity surface-emitting laser (VCSEL) arrays can be designed to operate in in-phase mode. However, the nonuniformities in fabrication process impact the resonance selection and the devices do not follow expected trends. Coherent coupling was demonstrated in three-element VCSEL arrays via phase tuning of elements. In-phase mode and out-of-phase mode were both achieved in most of the arrays. Moreover, coherent coupling can decrease the threshold current of elements in the array. Improved output power was also clearly observed when the array operated in the in-phase mode. Arbitrary phase combination of the array elements can be obtained via the phase tuning. This technology is able to improve the reproducibility and practicability of the implant-defined coherently coupled VCSEL array.

Background: Bovine leptospirosis is a widespread zoonotic disease, leading to serious economic losses in animal production and causing potential hazards to human health. Leptospiral lipopolysaccharide (L-LPS) plays an important role in leptospirosis pathogenicity., Objectives: With respect to L-LPS endotoxin-like activity, we examined bovine immune response to L-LPS and the inhibitory ability of bovine myeloid antimicrobial peptide-28 (BMAP-28) against L-LPS-induced immune activation in bovine cells., Materials and Methods: In this study, L-LPS-induced proinflammatory cytokine production in bovine cells was quantitatively measured with real-time PCR and ELISA, and we determined which cell membrane receptors (toll-like receptor [TLR]2 and TLR4) played a major role. In addition, the ability of BMAP-28 to inhibit L-LPS-induced endotoxin-like immune activation in bovine cells was determined by the decrease in cytokine secretion., Results: L-LPS showed the ability to induce cytokine production in bovine cells, and its induction was TLR2-dependent. BMAP-28 was used to inhibit L-LPS-induced endotoxin-like activity. The function of BMAP-28 was to inhibit LPS-induced TLR2 expression and cytokine production., Conclusions: In this study, the L-LPS immune response of bovine cells was significant, indicating that TLR2 is the predominant receptor for L-LPS. Due to L-LPS endotoxin-like activity, we found a strategy through using BMAP-28 to prevent L-LPS-induced TLR2-dependent immune activation in bovine cells.

In-phase coherently coupled vertical cavity surface emitting laser (VCSEL) hexagonal arrays were fabricated using proton implantation. The near-field profiles, far-field profiles, and emission spectra under different injection currents were tested and analyzed. The arrays can maintain in-phase single mode in a considerably wide current range from 10 mA (I(th)) to 35 mA (3.5×I(th)), exhibiting excellent beam quality. The far-field divergence angle of the in-phase coupled array is 2.5 degrees. Approximately 29% of total power is localized in the central lobe. Compared with square structure arrays, hexagonal arrays can maintain a more stable in-phase mode because of stronger coupling among the elements. The maximum output power of 4.9 mW was obtained under pulse wave condition. The simulation of far-field was carried out to match the in-phase operation test results. The performance enhancement of the array is attainable if the condition of heat dissipation is better. The process procedure of proton implantation is relatively simple and of low cost. It can be used as an alternative to coherently coupled array implementations.

Background: Enterovirus 71 (EV71) is one of the major etiological pathogens of hand, foot and mouth disease (HFMD) and can cause severe cerebral and pulmonary complications and even fatality. MicroRNAs (miRNAs), a class of small non-coding RNA molecules, play an important role in post-transcriptional regulation of gene expression and thereby influencing various physiological and pathological processes. Increasing evidence suggests that miRNAs act as key effector molecules in the complicated pathogen-host interactions. However, the roles of miRNAs in EV71 infection and pathogenesis are not well understood., Methods: To identify special miRNAs involved in EV71 infection, a microarray assay was performed to study the expression pattern of miRNAs in EV71-infected human rhabdomyosarcoma cells (RD cells) and uninfected RD cells. We further predicted the putative target genes for the dysregulated miRNAs using the online bioinformatic algorithms (TargetScan, miRanda and PicTar) and carried out functional annotation including GO enrichment and KEGG pathway analysis for miRNA predicted targets. Then, the results of microarray were further confirmed by quantitative RT-PCR., Results: Totally, 45 differentially expressed miRNAs ware identified by microarray, among which 36 miRNAs were up-regulated and 9 were down-regulated. 7166 predicted target genes for the dysregulated miRNAs were revealed by using TargetScan in conjunction with miRanda and PicTar. The GO annotation suggested that predicted targets of miRNAs were enriched into the category of signal transduction, regulation of transcription, metabolic process, protein phosphorylation, apoptotic process and immune response. KEGG pathway analysis suggested that these predicted target genes were involved in many important pathways, mainly including endocytosis and focal adhesion, MAPK signaling pathway, hypertrophic cardiomyopathy, melanogenesis and ErbB signaling pathway. The expression levels of 8 most differentially up-regulated miRNAs and 3 most differentially down-regulated miRNAs were confirmed by qRT-PCR. The expressions of hsa-miR-4530, hsa-miR-4492, hsa-miR-6125, hsa-miR-494-3p, hsa-miR-638, hsa-miR-6743-5p, hsa-miR-4459 and hsa-miR-4443 detected by qRT-PCR were consistent with the microarray data., Conclusion: These results might extend our understanding to the regulatory mechanism of miRNAs underlying the pathogenesis of EV71 infection, thus strengthening the preventative and therapeutic strategies of HFMD caused by EV71.

The two-dimensional (2D) triangle lattice air hole photonic crystal (PC) GaN-based light-emitting diodes (LED) with double-layer graphene transparent electrodes (DGTE) have been produced. The current spreading effect of the double-layer graphene (GR) on the surface of the PC structure of the LED has been researched. Specially, we found that the part of the graphene suspending over the air hole of the PC structure was of much higher conductivity, which reduced the average sheet resistance of the graphene transparent conducting electrode and improved the current spreading of the PC LED. Therefore, the work voltage of the DGTE-PC LED was obviously decreased, and the output power was greatly enhanced. The COMSOL software was used to simulate the current density distribution of the samples. The results show that the etching of PC structure results in the degradation of the current spreading and that the graphene transparent conducting electrode can offer an uniform current spreading in the DGTE-PC LED., Pacs: 85.60.Jb; 68.65.Pq; 42.70.Qs.

Introduction: Stenotrophomonas maltophilia infections have recently increased in importance in China, particularly in intensive care units (ICUs). The aim of this study was to investigate the clinical characteristics and risk factors of S. maltophilia infection in ICU of a hospital in northwest China., Methodology: The characteristics and outcomes of patients with any type of S. maltophilia infection at Shaanxi Provincial People's Hospital, Shaanxi, China, over a two-year period (from July 2011 to June 2013) were studied. S. maltophilia antimicrobial susceptibility was tested with the agar dilution method. The risk factors for all-cause in-hospital mortality were assessed with multivariate logistic regression., Results: Forty patients (median age, 72 years; 77.5% males) with S. maltophilia infection were identified. The main type of infection was lower respiratory tract infection (97.5%); one patient had a bloodstream infection. A total of 97.5% patients were infected with two or more organisms at the same time. The main characteristics of the patients were prolonged use of mechanical ventilation, urethral catheter, and central venous catheter before the infections occurred. The case number of infection was not different in the four seasons. High in vitro sensitivity was observed to minocycline (91.2%), levofloxacin (85.3%), and trimethoprim-sulfamethoxazole (79.4%). Most patients received therapy with a combination of agents. The crude mortality was 50%. By multivariate analysis, low albumin content and hypotension were the independent prognostic factors for mortality., Conclusions: Appropriate antimicrobial treatment had no positive impact on mortality. The impacts of albumin supplements and increasing blood pressure on mortality require further clinical studies.

Aim: To investigate cytokeratin 8 (CK8) overexpression during hepatitis C virus (HCV) infection and its pathogenesis, and the effect of ectopic CK8 expression on hepatoma cell lines., Methods: We successfully established an in vitro HCV cell culture system (HCVcc) to investigate the different expression profiles of CK8 in Huh-7-HCV and Huh-7.5-HCV cells. The expression of CK8 at the mRNA level was determined by real-time polymerase chain reaction (RT-PCR). The expression of CK8 at the protein level was evaluated by Western blotting. We then constructed a eukaryotic expression combination vector containing the coding sequence of human full length CK8 gene. CK8 cDNA was amplified by reverse transcription-PCR and inserted into pEGFP-C1 and the positive clone pEGFP-CK8 was obtained. After confirming the sequence, the recombinant plasmid was transfected into SMMC7721 cells with lipofectamine2000 and CK8 expression was detected using inverted fluorescence microscopy, RT-PCR and Western blotting. Besides, we identified biological function of CK8 on SMMC7721 cells, including cell proliferation, cell cycle and apoptosis detection., Results: RT-PCR showed that the expression level of CK8 in Huh-7-HCV and Huh-7.5-HCV cells was 2.88 and 2.95 times higher than in control cells. Western blot showed that CK8 expression in Huh-7-HCV and Huh-7.5-HCV cells was 2.53 and 3.26 times higher than that in control cells, respectively. We found that CK8 at mRNA and protein levels were both significantly increased in HCVcc. CK8 was up-regulated in SMMC7721 cells. CK8 expression at the mRNA level was significantly upregulated in SMMC7721/pEGFP-CK8 cells. CK8 expression in SMMC7721/ pEGFP-CK8 cells was 2.69 times higher than in SMMC7721 cells, and was 2.64 times higher than in SMMC7721/pEGFP-C1 cells. CK8 expression at the protein level in SMMC7721/pEGFP-CK8 cells was 2.46 times higher than in SMMC7721 cells, and was 2.29 times higher than in SMMC7721/pEGFP-C1 cells. Further analysis demonstrated that forced expression of CK8 slowed cell growth and induced apoptosis of SMMC7721 cells., Conclusion: CK8 up-regulation might have a functional role in HCV infection and pathogenesis, and could be a promising target for the treatment of HCV infection.

Cooperation or interaction of programmed cell death-1 (PD-1) and T cell immunoglobulin and mucin domain-containing molecule-3 (Tim-3) molecules is more relevant than either molecule alone to immune dysfunction in chronic viral infection and cancers. This study simultaneously investigated polymorphisms at PD1 +8669 and TIM3 -1516 loci in 845 hepatitis B virus (HBV) chronically infected patients [151 asymptomatic carriers, 202 chronic hepatitis, 221 cirrhosis and 271 hepatocellular carcinoma (HCC)], 141 HBV infection resolvers and 318 healthy controls. Multivariate analysis showed that, in addition to gender, age, ALT, albumin and HBV DNA, PD1 +8669 genotype AA was associated with cirrhosis compared with patients without cirrhosis (OR, 2.410; P=0.001). TIM3 -1516 genotypes GT+TT, together with gender, age, ALT, AST, direct bilirubin, albumin and HBeAg status, were associated with HCC compared with cirrhosis patients without HCC (OR, 2.142; P=0.011). The combined carriage of PD1 +8669 AA/TIM3 -1516 GT or TT was higher in cirrhosis and HCC pooled patients than in patients without cirrhosis (OR, 2.326; P=0.020) and in HCC patients than in cirrhosis patients (OR, 2.232; P=0.013). These data suggest that PD1 and TIM3 polymorphisms may differentially and interactively predispose cirrhosis and HCC in chronic HBV infection., (Copyright © 2012 Elsevier B.V. All rights reserved.)

To identify sociodemographic characteristics and risk factor of Demodex infestation, 756 students aged 13-22 years in Xi'an, China were sampled for the school-based cross-sectional study. Demodex was examined using the cellophane tape method (CTP). The results showed that the total detection rate of Demodex was 67.6%. Logistic regression analysis revealed that five variables (gender, residence, sharing sanitary ware, frequency of face-wash per day, and use of facial cleanser) were found to be uncorrelated with Demodex infestation, whereas three variables (age, skin type, and skin disease) were found to be independent correlates. Students aged over 18 years had 22.1 times higher odds of Demodex infestation compared to those under 16 years and students aged 16-18 years also had 2.1 times higher odds compared to those aged 13-15 years. Odds of having a Demodex infestation for oily or mixed skin were 2.1 times those for dry or neutral skin. Students with a facial skin disease had 3.0 times higher odds of being infested with Demodex compared to those without. The inception rate of students with facial dermatoses increased in parallel with increasing mite count. The inception rates were 21.3%, 40.7%, 59.2%, and 67.7% in the negative, mild, moderate, and severe infestation groups, respectively (χ(2)=60.6, P<0.001). Specifically, the amount of infested mites and inception rate of acne vulgaris were positively correlated (R(2)=0.57, moderate infestation odds ratio (OR)=7.1, severe infestation OR=10.3). It was concluded that Demodex prevalence increases with age, and Demodex presents in nearly all adult human. Sebaceous hyperplasia with oily or mixed skin seems to favour Demodex proliferation. Demodex infestation could be associated with acne vulgaris. The CTP is a good sampling method for studies of Demodex prevalence.

Background: Currently, up-regulated proteins and apoptosis in hepatitis C is a hot topic in exploring the pathogenic mechanism of Heptitis C Virus(HCV). Some recent studies shows that prohibitin is overexpressed in cells expressing HCV core proteins, and up-regulated prohibitin is also found in human hepatoma cell line HCC-M, lung cancer, prostate cancer, and other cancers. Prohibitin is an important member of the membrane protein superfamily, and it plays a role of molecular chaperones in mitochondrial protein stability. Meanwhile, it has a permissive action on tumor growth or acts as an oncosuppressor. Based on our previously established the in vitro HCV cell-culture system (HCVcc), here we aimed to investigate the different expression profiles of prohibitin in Huh-7-HCV and Huh-7.5-HCV cells, Methods: The total cellular RNA of Huh-7, Huh-7.5, Huh-7-HCV and Huh-7.5-HCV cells were extracted, and then the first-strand cDNA was reversely transcribed. The expression of prohibitin at the mRNA level was assessed by real-time PCR with GAPDH as the control. Furthermore, the expression of prohibitin at the protein level was evaluated by western blot with GAPDH as an internal control., Results: Our results of real-time PCR showed that the mRNA expression level of prohibitin in Huh-7-HCV cells was 2.09 times higher than that in Huh-7 cells, while, the mRNA level of prohibitin in Huh-7.5-HCV cells was 2.25 times higher than that in Huh-7.5 cells. The results of western blot showed that the protein expression level of prohibitin in Huh-7-HCV cells was 2.38 times higher than that in Huh-7 cells, while the protein expression of prohibitin in Huh-7.5-HCV cells was 2.29 times higher than that in Huh-7.5 cells., Conclusions: The expression of prohibitin was relatively high in Huh-7-HCV and Huh-7.5-HCV cells harboring in vitro transcribed full-length HCV RNA.

Aim: To construct an eukaryotic expression vector containing the coding region of human full length cytokeratin 8 gene and to detect its expression in SMMC7721 cells., Methods: CK8 cDNA was amplified by RT-PCR and cloned to pMD18-T simple vector. After confirming the sequence, the cDNA was inserted into pEGFP-C1 and the positive clone pEGFP-CK8 was obtained. The recombinant plasmid was transfected into SMMC7721 cells with Lipofectamine(TM);2000 and the expression was detected by fluorescence microscope, real time PCR and Western blot. The physical-chemical properties, signal peptide and functional motifs were predicted by the bioinformatics software., Results: PCR, restriction enzyme digestion and DNA sequencing showed that the recombinant plasmid contained the coding region of full length CK8 gene. Observation under fluorescence microscope and the results of real time PCR and western blot indicated CK8 was over-expressed in SMMC7721 cells., Conclusion: The eukaryotic expression vector containing the CK8 gene was successfully constructed and expressed, which provides a basis for the study for biological function of CK8.

Aim: To clone and express human keratin 8 gene cDNA in E.coli., Methods: Human cytokeratin 8 gene cDNA was amplified by RT-PCR from genomic RNA of human cell line 7721. The amplified cytokeratin 8 gene cDNA was cloned into pMD18-T vector. Then, the CK8 cDNA was amplified by PCR from recombinant plasmid pMD18-CK8, and was subcloned into pET-28a(+) expression vector. The recombinant plasmid pET-28a-CK8 DNA was transformed into E.coli DH5alpha strain., Results: Human cytokeratin 8 gene cDNA was cloned, and the recombinant plasmid pMD18-CK8 was transformed into E.coli. The CK8 cDNA was subcloned into E.coli DH5alpha strain, and successfully expressed in E.coli., Conclusion: Human cytokeratin 8 gene cDNA is cloned, and successfully expressed in E.coli, which lay the foundation of further study on the CK8 biological properties and functions.

Aim: The present study examined the differential expression of proteins in HuH-7 cells and HuH-7 cells harboring in vitro-transcribed full-length hepatitis C virus 1b RNA (HuH-7-HCV), and elucidated the cellular responses to HCV replication., Methods: The protein profiles of matched pairs of HuH-7-HCV cells and HuH-7 mock cells were analyzed by 2-D electrophoresis (2DE). Solubilized proteins were separated in the first dimension by isoelectric focusing, and by 12.5% SDS-PAGE in the second dimension. The differential protein expression was analyzed by use of image analysis software to identify candidates for HCV infection-associated proteins., Results: In total, 29 protein spots showed increases and 25 protein spots showed decreases in signal in HuH-7-HCV cell 2DE profiles as compared with HuH-7 mock cells. In the next step, the 10 spots showing the greatest increase and the 10 spots showing the greatest decrease were excised from gels and the proteins present were identified by Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometer (MALDI-TOF MS) or MALDI-TOF/TOF MS. In total, 13 proteins were identified successfully. The potential significance of the differential expression due to HCV replication was discussed., Conclusion: Our study identifies changes in the proteome of HuH-7 cells in the presence of HCV replication and yields information of the mechanism of HCV pathogenesis. These results will be useful for the identification of HCV infection-associated proteins that could be molecular targets for treatment.

Background/aims: Higher virological and biochemical response rates were obtained in many clinical trials in Caucasian patients with chronic hepatitis C (CHC) after treating with peginterferon than in those with interferon. However, it is not clear whether this conclusion can be extrapolated to patients with Chinese ethnic origin and which type of peginterferon or interferon was more effective in treating Chinese CHC patients? The aim of this study was to perform a meta-analysis of randomized controlled trials (RCT) and compare peginterferon with interferon in treating Chinese patients with CHC., Methodology: The outcome measures were virological response (end-of-treatment virological response (ETVR) and sustained virological response (SVR)). Biochemical response (end-of-treatment biochemical response (ETBR) and sustained biochemical response (SBR) were also introduced as supplement to the evaluation on effect of peginterferon and interferon in treating Chinese patients with CHC. The measure of association employed was relative risk (RR) calculated by the random-effect model, with heterogeneity, sensitivity and subgroup analyses., Results: Of the 110 studies screened, 5 RCTs including 499 patients (peginterferon 258, interferon 241) were analyzed. SVR and SBR obtained in patients treated with peginterferon were significantly higher than patients treated with interferon (60% vs. 35.53%, RR, 1.95; 95% confidence interval (CI), 1.17-3.24; p<0.05; 57.30% vs. 40.38%, RR, 1.36; 95% CI, 1.08-1.71; p<0.05), but the difference is not evident in ETVR and ETBR. In the subgroup analyses, ETVR and SVR obtained in patients treated with peginterferon alpha-2a (79.75% and 48.1%) were significantly higher than with interferon alpha-2a (35.37% vs. 19.05%, RR, 2.51; 95% CI, 1.74-3.63; p<0.05), but the difference is not evident between peginterferon alpha-2b and interferon alpha-2b. SBR obtained with peginterferon was also significantly higher than with interferon (57.3% vs. 40.38%, RR, 1.36; 95% CI, 1.08-1.71; p<0.05), but the difference is not evident in EBVR. Higher SVR were obtained in genotype 1 Chinese patients treated with peginterferon compared with those treated with interferon (48.45% vs. 23.24%, RR, 2.00; 95% CI, 1.48-2.7; p<0.01), but the ETVR were similar. The effects of peginterferon and interferon in genotype non-1 patients were also similar. Higher ETVR and SVR were obtained in patients treated with peginterferon alpha-2b compared with those treated with peginterferon alpha-2a (97.56% vs. 79.75% and 82.93% vs. 56.96%; p<0.05 and p<0.01, respectively). SVR obtained in patients treated with peginterferon alpha-2b were significantly higher than those treated with peginterferon alpha-2a (73.53% vs. 41.73%, p<0.01) in treating genotype 1 Chinese patients, but the superiority didn't appear in treating genotype non-1 patients., Conclusions: Both peginterferon alpha-2a and peginterferon alpha-2b might be recommended for Chinese ethnic original patients with CHC. Peginterferon alpha-2b might be more effective, with an expected relative higher virological response and biochemical response, in treating Chinese CHC patients.

Objective: To investigate the effects of hepatitis C virus (HCV) on transcription regulation genes of host cells by gene chip assays in cultured cells with intact HCV genome., Methods: Huh-7 hepatoma cells were cultured and infected with in vitro constructed HCV. The total RNAs, proteins and cell culture supernatants of HCV infected cells and control cells were isolated. Proteins and cell culture supernatants were used to detect the HCV replication and protein expression in cell culture system. The HCV protein expression was detected with Western blotting. Released HCV from infected cells was analyzed by real-time fluorescence quantitative PCR. Total RNA was qualified using 10 g/L agarose gel electrophoresis. cRNA was synthesized, fluorescence labeled and purified, then hybridized with Agilent oligo microarray (20173 probes). Differential expression of genes related to transcription in cell culture system was analyzed., Result: HCV was positive in cell culture supernatants and HCV protein expression was also positive according to Western blotting results. Eleven up-regulated and 11 down-regulated genes related to transcription were found after Agilent gene chip screening., Conclusion: Intact hepatitis C virus cell culture system provides an useful tool for study on the affects of HCV infection on transcription regulation genes in host cells.

Objective: The intact hepatitis C virus (HCV) cell culture system has provided a powerful tool for studying the interaction between HCV and host cell. We applied proteomic techniques to globally analyze the protein expression profiles of Huh7 in the absence and presence of HCV, with the aim to elucidate the host cell components response to HCV., Methods: Proteomic and molecular biology techniques were used for this aim., Results: The expression of many proteins, including regucalcin, centriolin and several keratins, was up-regulated, after HCV transfection. Among the down-regulated expression proteins, the heat-shock protein family was of prime significance. Subsequently, we studied the role of heat stress in the interaction between HCV and host cell, based on proteomics analysis. We found heat stress did not affect the production of HCV in this in vitro cell culture system., Conclusions: Moreover, this study also provided the global information of proteomic alteration of Huh7 cells in the presence of intact HCV and further improved the understanding of the mechanism of interaction between HCV and host cell. We also draw a conclusion that heat stress had no affect on the production of HCV., (Copyright 2008 S. Karger AG, Basel.)

Lack of proper study models has brought difficulties in the study of the mechanism of viral infection, life cycle and pathogenic mechanism of hepatitis C virus (HCV) and also become the major obstacles in development of efficient vaccine and new drugs for hepatitis C. In recent years, the establishment of robust HCV cell culture infection system and HCV transgenic animal provide powerful tools for the analysis of host virus interactions, which facilitate the discovery of antiviral drugs and vaccines for this important human pathogen.

Aim: To construct the prokaryotic expression plasmid of hWAPL, induce the expression of the protein and prepare polyclonal antibody., Methods: The hWAPL cDNA was amplified by RT-PCR from HeLa cells derived RNA and then cloned to pMD18-T according to A-T. The DNA fragment was isolated and linked to prokaryotic expression vector pET28a. The recombinant protein was induced by IPTG and identified by SDS-PAGE. Then it was injected into mice to prepare polyclonal antibody. The immunogenicity and specificity of the recombinant protein were identified by ELISA and Western blot., Results: The sequencing, PCR and endonucleases digestion results showed that the hWAPL fragment was correctly inserted into pMD18-T and pET28a vectors. SDS-PAGE showed 25,000 fusion hWAPL protein was expressed in BL21 cells. The titer of polyclonal antibody was 1:3 200 by indirect ELISA. The 25,000 fusion hWAPL protein was detected by SDS-PAGE and Western blot., Conclusion: The hWAPL protein can be expressed by prokaryotic vector pET28a. Furthermore, the expression protein can be used as immunogen to generate antibodies with specificity.