Search

Introduction Potency is a critical quality attribute for controlling quality consistency and relevant biological properties of vaccines. Owing to the high demand for animals, lengthy operations and high variability of in vivo methods, in vitro alternatives for human vaccine potency assays are extensively developed. Areas covered Herein, in vivo and in vitro methods for potency assays of previously licensed human vaccines were sorted, followed by a brief description of the background for substituting in vivo methods with in vitro alternatives. Based on the analysis of current research on the substitution of vaccine potency assays, barriers and suggestions for substituting were proposed. Expert opinion Owing to the variability of in vivo methods, the correlation between in vivo and in vitro methods may be low. One or more in vitro method(s) that determine the vaccine antigen content and functions, should be established. Since the substitution involves with the change of critical quality attributes and specifications, the specifications of in vitro methods should be appropriately set to maintain the efficacy of vaccines. For novel vaccines in research and development, in vitro methods for monitoring the consistency and relevant biological properties, should be established based on reflecting the immunogenicity of vaccines.

Abstract Recombinant proteins are gaining increasing popularity for treating human diseases. The clinical effectiveness of recombinant proteins is directly related to their biological activity, which is an important indicator in drug development and quality control. However, certain recombinant proteins have unclear or complex signal pathways, making detecting their activity in vitro difficult. For instance, recombinant human endostatin (endostatin), a new antitumor drug developed in China, lacks a sensitive and stable assay for its biological activity since being market approval. To address this issue, we performed a genome‐wide screening of immortalized human umbilical vein endothelial cells (HUVECs) using a CRISPR/Cas9 knockout library containing 20,000 targeted genes. We identified two potential endostatin‐resistant genes, NEPSPP and UTS2, and successfully constructed a highly sensitive cell line, HUVEC‐UTS2‐3#, by knocking down the UTS2 gene. Based on the optimized parameters of HUVEC‐UTS2‐3# cells, we established a new method for detecting the biological activity of endostatin. The method was validated, and it produced results consistent with primary HUVEC cells but with higher sensitivity and more stable data. The use of gene‐editing technology provides a novel solution for detecting the biological activity of recombinant proteins that other methods cannot detect.

Recombinant adeno-associated virus (rAAV) has emerged as a prominent vector for in vivo gene therapy, owing to its distinct advantages. Accurate determination of the rAAV genome titer is crucial for ensuring the safe and effective administration of clinical doses. The evolution of the rAAV genome titer assay from quantitative PCR (qPCR) to digital PCR (dPCR) has enhanced accuracy and precision, yet practical challenges persist. This study systematically investigated the impact of various operational factors on genome titration in a single-factor manner, aiming to address potential sources of variability in the quantitative determination process. Our findings revealed that a pretreatment procedure without genome extraction exhibits superior precision compared with titration with genome extraction. Additionally, notable variations in titration results across different brands of dPCR instruments were documented, with relative standard deviation (RSD) reaching 23.47% for AAV5 and 11.57% for AAV8. Notably, optimal operations about DNase I digestion were identified; we thought treatment time exceeding 30 min was necessary, and there was no need for thermal inactivation after digestion. And we highlighted that thermal capsid disruption before serial dilution substantially affected AAV genome titers, causing a greater than ten-fold decrease. Conversely, this study found that additive components of dilution buffer are not significant contributors to titration variations. Furthermore, we found that repeated freeze–thaw cycles significantly compromised AAV genome titers. In conclusion, a comprehensive dPCR titration protocol, incorporating insights from these impact factors, was proposed and successfully tested across multiple serotypes of AAV. The results demonstrate acceptable variations, with the RSD consistently below 5.00% for all tested AAV samples. This study provides valuable insights to reduce variability and improve the reproducibility of AAV genome titration using dPCR.

The kinematic information of the Radcliffe wave (RW) is essential for determining its existence and gaining insights into its origin and evolution. In this work, we present an accurate measurement of the vertical velocity ( V _Z ) of the RW by incorporating the radial velocity (RV) measures through two methods, which is crucial but was neglected previously. First, the velocities are measured toward young stars, using their RV measurements from APOGEE-2 and proper motion measurements from Gaia DR3. Second, we combine RV measurements toward clouds with proper motion measurements of associated young stellar objects to determine the vertical velocities of the clouds. The results reveal that the oscillations in V _Z are not synchronous with the vertical coordinate Z , which differs from the conclusions of previous studies. Instead, we find a 5 km s ^−1 kpc ^−1 gradient in V _Z along the RW, exhibiting a dipole-like pattern. Consequently, the kinematic arrangement does not show a corresponding coherence with the spatial arrangement, bringing the RW model into question.

C _4 H and c -C _3 H _2 , as unsaturated hydrocarbon molecules, are important for forming large organic molecules in the interstellar medium. We present mapping observations of C _4 H ( N = 9−8) lines, c -C _3 H _2 ( J _Ka _, _Kb = 2 _1,2 –1 _0,1 ), and H ^13 CO ^+ ( J = 1−0) toward 19 nearby cold molecular cores in the Milky Way with the IRAM 30 m telescope. C _4 H 9–8 was detected in 13 sources, while c -C _3 H _2 was detected in 18 sources. The widely existing C _4 H and c -C _3 H _2 molecules in cold cores provide material to form large organic molecules. Different spatial distributions between C _4 H 9–8 and c -C _3 H _2 2–1 were found. The relative abundances of these three molecules were obtained under the assumption of local thermodynamic equilibrium conditions with a fixed excitation temperature. The abundance ratio of C _4 H to c -C _3 H _2 ranged from 0.34 ± 0.09 in G032.93+02 to 4.65 ± 0.50 in G008.67+22. A weak correlation between C _4 H/H ^13 CO ^+ and c -C _3 H _2 /H ^13 CO ^+ abundance ratios was found, with a correlation coefficient of 0.46, which indicates that there is no tight astrochemical connection between C _4 H and c -C _3 H _2 molecules.

Dense outflowing gas, traced by transitions of molecules with large dipole moment, is important for understanding mass loss and feedback of massive star formation. HCN 3-2 and HCO ^+ 3-2 are good tracers of dense outflowing molecular gas, which are closely related to active star formation. In this study, we present on-the-fly mapping observations of HCN 3-2 and HCO ^+ 3-2 toward a sample of 33 massive star-forming regions using the 10-m Submillimeter Telescope. With the spatial distribution of line wings of HCO ^+ 3-2 and HCN 3-2, outflows are detected in 25 sources, resulting in a detection rate of 76%. The optically thin H ^13 CN and H ^13 CO ^+ 3-2 lines are used to identify line wings as outflows and estimate core mass. The mass M _out , momentum P _out , kinetic energy E _K , force F _out , and mass-loss rate ${\dot{M}}_{{\rm{out}}}$ of outflow and core mass, are obtained for each source. A sublinear tight correlation is found between the mass of dense molecular outflow and core mass, with an index of ∼0.8 and a correlation coefficient of 0.88.

Deuterated molecules are valuable probes for investigating the evolution and the kinematics in the earliest stages of star formation. In this study, we conduct a comprehensive investigation by performing a single-point survey of 101 starless clump candidates, and carrying out on-the-fly (OTF) observations of 11 selected sources, focusing on deuterated molecular lines using the IRAM 30 m telescope. In the single-point observation, we make 46 detections for DCO ^+ J = 1−0, 12 for DCN J = 1−0, 51 for DNC J = 1−0, 7 for N _2 D ^+ J = 1−0, 20 for DCO ^+ J = 2−1, and 10 for DCN J = 2−1. The starless clump candidates with deuterated molecule detections exhibit lower median kinetic temperatures and a narrower H _2 CO (1 _(0,1) −0 _(0,0) ) median full width at half maximum compared to those without such detections, while simultaneously displaying similar median values of 1.1 mm intensity, mass, and distance. Furthermore, our OTF observations reveal that deuterated molecules predominantly have peaks near the 1.1 mm continuum peaks, with the DCO ^+ J = 1−0 emission demonstrating higher intensity in the deuterated peak region compared to the DCN and DNC J = 1−0 emissions. Additionally, the majority of emissions from deuterated molecules and ^13 C isotopologues exhibit peak positions close to those of the 1.1 mm continuum peaks. By analyzing the 20″ × 20″ regions with strongest deuterated emissions in the OTF observations, we estimated deuterated abundances of 0.004−0.045, 0.011−0.040, and 0.004−0.038 for D _frac (HCN), D _frac (HCO ^+ ), and D _frac (HNC), respectively. The differential detection of deuterated molecular lines in our OTF observations could be attributed to variations in critical densities and formation pathways.

Peptide-like molecules, which have a close connection with the origin of life, have been detected in the Universe. Mapping observations of HCONH _2 and CH _3 CONH _2 , two of the simplest peptide-like molecules, are performed toward the Sagittarius B2 (Sgr B2) complex with the IRAM 30 m telescope. Seven transitions of HCONH _2 and five transitions of CH _3 CONH _2 are used in the analysis. The spatial distributions of the excitation temperature and column density of HCONH _2 in the molecular envelope of Sgr B2 are obtained by rotation diagrams. Assuming the same excitation temperature of HCONH _2 , the column densities of CH _3 CONH _2 are also calculated. The results show that the excitation temperature ranges from 6 to 46 K in the molecular envelope of Sgr B2. The abundance ratios between HCONH _2 and CH _3 CONH _2 are calculated to explore the relationship between them, as are those between HCONH _2 and HNCO. The abundance ratio of CH _3 CONH _2 /HCONH _2 varies from 10% to 20%, while that of HCONH _2 /HNCO ranges from 1.5% to 10%. CH _3 CONH _2 is enhanced with respect to HCONH _2 in the northwest region of Sgr B2. One transition of H ^13 CONH _2 is detected toward 12 positions of Sgr B2, from which a ^12 C/ ^13 C ratio of 28.7 is obtained. A time-dependent chemical model with a short-duration X-ray burst is used to explain the observed abundances of HCONH _2 and CH _3 CONH _2 , with the best-fitting result at T _dust = 53–56 K. More chemical reactions are required to be included in the model since the modeled abundance is lower than the observed one at the observed T _dust .

Of all of the components in SARS-CoV-2 inactivated vaccines, nucleocapsid protein (N) is the most abundant and highly conserved protein. However, the function of N in these vaccines, especially its influence on the targeted spike protein’s response, remains unknown. In this study, the immunization of mice with the N protein alone was shown to reduce the viral load, alleviating pulmonary pathological lesions after challenge with the SARS-CoV-2 virus. In addition, co-immunization and pre-immunization with N were found to induce higher S-specific antibody titers rather than compromise them. Remarkably, the same trend was also observed when N was administered as the booster dose after whole inactivated virus vaccination. N-specific IFN-γ-secreting T cell response was detected in all groups and exhibited a certain relationship with S-specific IgG antibody improvements. Together, these data indicate that N has an independent role in vaccine-induced protection and improves the S-specific antibody response to inactivated vaccines, revealing that an interplay mechanism may exist in the immune responses to complex virus components.

Recombinant human interferon α2b (rhIFNα2b) is widely used as an antiviral therapy agent for the treatment of hepatitis B and hepatitis C. The current identification test for rhIFNα2b is complex. In this study, an anti-rhIFNα2b nanobody was discovered and used for the development of a rapid lateral flow strip for the identification of rhIFNα2b. RhIFNα2b was used to immunize an alpaca, which established a phage nanobody library. After five steps of enrichment, the nanobody I22, which specifically bound rhIFNα2b, was isolated and inserted into the prokaryotic expression vector pET28a. After subsequent purification, the physicochemical properties of the nanobody were determined. A semiquantitative detection and rapid identification assay of rhIFNα2b was developed using this novel nanobody. To develop a rapid test, the nanobody I22 was coupled with a colloidal gold to produce lateral-flow test strips. The developed rhIFNα2b detection assay had a limit of detection of 1 μg/mL. The isolation of I22 and successful construction of a lateral-flow immunochromatographic test strip demonstrated the feasibility of performing ligand-binding assays on a lateral-flow test strip using recombinant protein products. The principle of this novel assay is generally applicable for the rapid testing of other commercial products, with a great potential for routine use in detecting counterfeit recombinant protein products.

Abstract Emerging SARS-CoV-2 variants and the gradually decreasing neutralizing antibodies over time post vaccination have led to an increase in incidents of breakthrough infection across the world. To investigate the potential protective effect of the recombinant protein subunit COVID-19 vaccine targeting receptor-binding domain (RBD) (PS-RBD) and whole inactivated virus particle vaccine (IV) against the variant strains, in this study, rhesus macaques were immunized with PS-RBD or IV vaccine, followed by a Beta variant (B.1.351) challenge. Although neutralizing activity against the Beta variant was reduced compared with that against the prototype, the decreased viral load in both upper and lower respiratory tracts, milder pathological changes, and downregulated inflammatory cytokine levels in lung tissues after challenge demonstrated that PS-RBD and IV still provided effective protection against the Beta variant in the macaque model. Furthermore, PS-RBD-induced macaque sera possessed general binding and neutralizing activity to Alpha, Beta, Delta, and Omicron variants in our study, though the neutralizing antibody (NAb) titers declined by varying degrees, demonstrating potential protection of PS-RBD against current circulating variants of concern (VOCs). Interestingly, although the IV vaccine-induced extremely low neutralizing antibody titers against the Beta variant, it still showed reduction for viral load and significantly alleviated pathological change. Other correlates of vaccine-induced protection (CoP) like antibody-dependent cellular cytotoxicity (ADCC) and immune memory were both confirmed to be existing in IV vaccinated group and possibly be involved in the protective mechanism.

Despite the widespread use of hydrogels, methods offering improved control over gelation mechanisms and patterning are still sought for. Here, the authors explore potentiostatic control in electrochemical-chemical-chemical reactions on chitosan, alginate and alginate/PEDOT composite systems which allows selection of covalent and ionic gelation mechanisms as well as the growth rate of the hydrogel.

Regulatory science is a discipline that uses comprehensive methods of natural science, social science, and humanities to provide support for administrative decision-making through the development of new tools, standards, and approaches to assess the safety, efficacy, quality, and performance of regulated products. During the pandemics induced by infectious diseases, such as H1N1 flu, severe acute respiratory syndrome (SARS), and Middle East respiratory syndrome (MERS), regulatory science strongly supported the development of drugs and vaccines to respond to the viruses. In particular, with the support of research on drug regulatory science, vaccines have played a major role in the prevention and control of coronavirus disease 2019 (COVID-19). This review summarizes the overall state of the vaccine industry, research and development (R&D) of COVID-19 vaccines in China, and the general state of regulatory science and supervision for vaccines in China. Further, this review highlights how regulatory science has promoted the R&D of Chinese COVID-19 vaccines, with analyses from the aspects of national-level planning, relevant laws and regulations, technical guidelines, quality control platforms, and post-marketing supervision. Ultimately, this review provides a reference for the formulation of a vaccine development strategy in response to the current pandemic and the field of vaccine development in the post-pandemic era, as well as guidance on how to better respond to emerging and recurring infectious diseases that may occur in the future.

Coxsackievirus A6 (CA6) is one of the major causative agents of herpangina and hand-foot-mouth disease (HFMD). Since 2008, CA6 has circulated widely around the world. Especially in Asia-Pacific region CA6 had even replaced enterovirus A71 (EV71) and coxsackievirus A16 (CA16) as the main prevalent strain of HFMD. In the recent 10 years, monovalent and multivalent vaccines against CA6 have been researched and developed by manufacturers from China, Korea, and the USA. The neutralizing antibody titer is a key indicator for accurately evaluating immunogenicity of vaccine. However, so far, the World Health Organization international standard for CA6 neutralizing antibody has not been available. In order to meet the needs of evaluating the immunogenicity of vaccines against CA6, the first Chinese national standard for CA6 neutralizing antibody was established, which was conducted to ensure that methods used to measure the neutralizing antibody titers against CA6 are accurate, reliable, and comparable. Three lyophilized candidate standards (29#, 39# and 44#) were produced with 0.40 ml/vial from plasma samples donated by healthy individuals. The collaborative study showed that the 29# candidate standard could effectively minimize the variability in neutralization titers between labs and across challenging viruses of different genotypes (A, D1, and D3). Therefore, the 29# candidate sample was established as the first Chinese national standard for CA6 neutralizing antibody test. This standard has good long-term stability and was assigned a potency of 150 units per milliliter (U/ml) of CA6 neutralizing antibody. It will contribute to ensure uniformity of potency or activity of vaccines and potentially therapeutic antibody preparations.

Abstract Integrating different types of vaccines into a singular immunization regimen is an effective and accessible approach to strengthen and broaden the immunogenicity of existing coronavirus disease 2019 (COVID‐19) vaccine candidates. To optimize the immunization strategy of the novel mRNA‐based vaccine and recombinant protein subunit vaccine that attracted much attention in COVID‐19 vaccine development, we evaluated the immunogenicity of different combined regimens with the mRNA vaccine (RNA‐RBD) and protein subunit vaccine (PS‐RBD) in mice. Compared with homologous immunization of RNA‐RBD or PS‐RBD, heterologous prime‐boost strategies for mRNA and protein subunit vaccines failed to simultaneously enhance neutralizing antibody (NAb) and Th1 cellular response in this study, showing modestly higher serum neutralizing activity and antibody‐dependent cell‐mediated cytotoxicity for “PS‐RBD prime, RNA‐RBD boost” and robust Th1 type cellular response for “RNA‐RBD prime, PS‐RBD boost”. Interestingly, immunizing the mice with the mixed formulation of the two aforementioned vaccines in various proportions further significantly enhanced the NAb responses against ancestral, Delta, and Omicron strains and manifested increased Th1‐type responses, suggesting that a mixed formulation of mRNA and protein vaccines might be a more prospective vaccination strategy. This study provides basic research data on the combined vaccination strategies of mRNA and protein‐based COVID‐19 vaccines.

The theory of groundwater flow systems proposed by Tóth, which was based on the two-dimensional profile flow field, has become the theoretical basis for studying basin-scale groundwater circulation.The stagnation points on the two-dimensional profile can be used to accurately divide the spatial distribution of groundwater flow systems.However, the spatial distribution of groundwater flow systems and whether stagnation points exist in the 3D domain remain unknown.In this study, two typical three-dimensional basins, namely foreland-like basins with undulations only in the x direction and dune-type basins with undulations in both x and y directions, are used as typical examples.By using a combination of cosine functions to characterize the undulating water table, analytical solutions of head distribution in the two basins are derived.Moreover, by employing the TECPLOT software, the spatial distribution of the 3D groundwater flow systems is clearly shown.It is found out that stagnation points can develop in two basins.The stagnation points in the foreland-like basin constitute a horizontally extending stagnation line, which controls the spatial distribution of four flow systems in the basin, while the stagnation points in the dune-type basin form an arc-shaped stagnation line, which controls the spatial distribution of six flow systems in the basin.By studying the distribution of stagnation line in 3D basins and successfully dividing flow systems by using the stagnation line, this study leads to a deepened understanding of groundwater circulation in 3D basins.

Groundwater age and groundwater residence time contain important information about groundwater circulation and evolutionary processes, and have been widely used in the study of groundwater circulation patterns in basins.In this paper, we simulated a three-stage flow system model through a multi-stage groundwater flow system demonstrator, and simulated the groundwater age distribution and groundwater residence time distribution based, and found that the bottom of the basin, the downstream of the regional flow, and the basin retention area responded the latest.The local flow system in the shallow part has relatively low concentration values after stabilization, the intermediate flow system is also relatively low compared to the regional flow system in the deep part, and the stagnant zone has relatively large concentration values due to salt accumulation.The groundwater age distribution curves are single-peaked, and the circulation time of the regional flow system is greater than that of the intermediate flow system than that of the local flow system.The residence time distribution monitored in the discharge zones shows that different levels of recharge will produce early, middle and late peaks, and the peaks correspond exactly to the level of the groundwater flow system.It can be judged from the peaks in the discharge zones that the groundwater is recharged from the local, intermediate or regional flow system, and thus the source of contaminants can be determined.The present research results have some significance for the evolution of groundwater circulation and the improvement of groundwater flow system theory.

Identifying the permeability structure of the dam foundation rock mass is important for the formulation of seepage control schemes. For the horizontally-stratified red-bed rock mass of the Yellow River Guxian Dam foundation, this paper adopted a "continuous, high-resolution" water pressure test data processing method to analyze the relationships between rock mass permeability and elevation, lithology and shear zones, clarifiy the permeability structure of the dam foundation rock mass, and propose seepage control advice. The results indicate that the Guxian Dam foundation rock mass is characterized by a structural and random permeability behavior. The structure shows that the permeability decreases with elevation. The randomness shows that the permeability randomly fluctuates due to lithology and shear zones. For the river-bed dam foundation rock mass, the bottom elevation of the weathering and unloading zone is 450 m or so, and the 1Lu bottom boundary is at the elevation of approximately 340 m. For the rock mass of the dam abutment, the 3Lu bottom boundary is located at the elevation of 560～580 m. The section of 350～360 m elevation has a certain thickness of soft rock, a poor development of shear structure, and a low permeability. It can be regarded as a confining layer and a potential target for the optimization of seepage control schemes. In view of the universal applicability, the method proposed in this paper can be used for reference in similar projects.

T cell immunoglobulin and ITIM domain (TIGIT) is a novel immune checkpoint that has been considered as a target in cancer immunotherapy. Current available bioassays for measuring the biological activity of therapeutic antibodies targeting TIGIT are restricted to mechanistic investigations because donor primary T cells are highly variable. Here, we designed a reporter gene assay comprising two cell lines, namely, CHO-CD112-CD3 scFv, which stably expresses CD112 (PVRL2, nectin-2) and a membrane-bound anti-CD3 single-chain fragment variable (scFv) as the target cell, and Jurkat-NFAT-TIGIT, which stably expresses TIGIT as well as the nuclear factor of activated T-cells (NFAT) response element-controlled luciferase gene, as the effector cell. The anti-CD3 scFv situated on the target cells activates Jurkat-NFAT-TIGIT cells through binding and crosslinking CD3 molecules of the effector cell, whereas interactions between CD112 and TIGIT prevent activation. The presence of anti-TIGIT mAbs disrupts their interaction, which in turn reverses the inactivation and luciferase expression. Optimization and validation studies have demonstrated that this assay is superior in terms of specificity, accuracy, linearity, and precision. In summary, this reliable and effective reporter gene assay may potentially be utilized in lot release control, stability assays, screening, and development of novel TIGIT-targeted therapeutic antibodies.

Aquifer diffusivity is a basic physical parameter used in hydrogeological calculations and is important for the evaluation and rational utilization of water resources, pollution prevention, and wetland protection. In this study, with the assumptions of aquifer isotropy, i.e., no vertical flow and constant saturated aquifer thickness, by using the Fourier transform and convolution theorem, a 1D analytical solution of the phase spectrum for an unconfined aquifer system was derived, which subsequently led to a phase spectrum solution for aquifer diffusivity. To test the efficacy, the proposed method was applied to a study site in the middle reach of the Hutuo River in the North China Plain. The estimated aquifer diffusivity ranged from 1.9 × 103 to 4.9 × 104 m2/d, with a mean of 2.2 × 104 m2/d, which was consistent with the results obtained using power spectral analysis, pumping tests, and inverse numerical models. The phase spectral approach proposed in this paper can estimate the aquifer properties on a larger scale. If long time series of hydraulic heads are available, it can estimate hydrogeological parameters accurately and quickly. Considering the similarity of the linearized governing equations, it can also be applied to the river–aquifer system and the confined aquifer system.

We previously screened 6 differentially expressed miRNAs in ovarian tissues of 4-vinylcyclohexene diepoxide (VCD)-treated premature ovarian failure (POF) model in SD rats, including miRNA-190a-5p, miRNA-98-5p, miRNA-29a-3p, miRNA-144-5p, miRNA-27b-3p, miRNA-151-5p. In this study, to investigate the mechanisms causing the onset of POF, we first identified miRNAs with earlier differential expression at consecutive time points in the VCD-treated rat POF model and explored the mechanisms by which the target miRNAs promote POF. The SD rats were injected with VCD for 15 days to induce POF. Additionally, we collected rat blood and ovaries at the same time every day for 15 consecutive days, and luteinizing hormone (LH), follicle-stimulating hormone (FSH), Anti-Mullerian hormone (AMH), and estradiol (E2) serum levels were detected by ELISA. Six miRNAs expression were measured in rat ovaries by qRT-PCR. Dual-luciferase reporter gene assays were employed to predict and verify the target gene (PHLPP1) of target miRNAs (miRNA-190a-5p). Western blot was examined to detect the expression levels of PHLPP1, AKT, p-AKT, FOXO3a, p-FOXO3a, and LHR proteins on the target gene PHLPP1 and its participation in the primordial follicular hyperactivation-related pathways (AKT-FOXO3a and AKT-LH/LHR). During the VCD modeling POF rat ovaries, miRNA-190a-5p was the first to show significant differential expression, i.e., 6th of VCD treating, and PHLPP1 was verified to be a direct downstream target of it. Starting from the 6th of VCD treatment, the more significant the up-regulation trend of miRNA-190a-5p expression, the more obvious the down-regulation trend of PHLPP1 and LHR mRNA and protein expression, accompanied by the more severe phosphorylation of AKT and FOXO3a proteins, thus continuously over-activating the rat primordial follicle to promote the development of POF. In conclusion, miRNA-190a-5p may become a potential biomarker for early screening of POF, and it can continuously activate primordial follicles in rats by targeting the expression of PHLPP1 and key proteins in the AKT-FOXO3a and AKT-LH/LHR pathways.

ABSTRACT We investigated the distribution, virulence, and pathogenic characteristics of mutated SARS-CoV-2 to clarify the association between virulence and the viral spreading ability of current and future circulating strains. Chinese rhesus macaques were infected with ancestral SARS-CoV-2 strain GD108 and Beta variant B.1.351 (B.1.351) and assessed for clinical signs, viral distribution, pathological changes, and pulmonary inflammation. We found that GD108 replicated more efficiently in the upper respiratory tract, whereas B.1.351 replicated more efficiently in the lower respiratory tract and lung tissue, implying a reduced viral shedding and spreading ability of B.1.351 compared with that of GD108. Importantly, B.1.351 caused more severe lung injury and dramatically elevated the level of inflammatory cytokines compared with those observed after infection with GD108. Moreover, both B.1.351 and GD108 induced spike-specific T-cell responses at an early stage of infection, with higher levels of interferon gamma (IFN-γ) and tumor necrosis factor alpha (TNF-α) in the B.1.351 group and higher levels of interleukin 17 (IL-17) in the GD108 group, indicating a divergent pattern in the T-cell-mediated inflammatory “cytokine storm.” This study provides a basis for exploring the pathogenesis of SARS-CoV-2 variants of concern (VOCs) and establishes an applicable animal model for evaluating the efficacy and safety of vaccines and drugs. IMPORTANCE One of the priorities of the current SARS-CoV-2 vaccine and drug research strategy is to determine the changes in transmission ability, virulence, and pathogenic characteristics of SARS-CoV-2 variants. In addition, nonhuman primates (NHPs) are suitable animal models for the study of the pathogenic characteristics of SARS-CoV-2 and could contribute to the understanding of pathogenicity and transmission mechanisms. As SARS-CoV-2 variants continually emerge and the viral biological characteristics change frequently, the establishment of NHP infection models for different VOCs is urgently needed. In the study, the virulence and tissue distribution of B.1.351 and GD108 were comprehensively studied in NHPs. We concluded that the B.1.351 strain was more virulent but exhibited less viral shedding than the latter. This study provides a basis for determining the pathogenic characteristics of SARS-CoV-2 and establishes an applicable animal model for evaluating the efficacy and safety of vaccines and drugs.

To cope with the decline in COVID-19 vaccine-induced immunity caused by emerging SARS-CoV-2 variants, a heterologous immunization regimen using chimpanzee adenovirus vectored vaccine expressing SARS-CoV-2 spike (ChAd-S) and an inactivated vaccine (IV) was tested in mice and non-human primates (NHPs). Heterologous regimen successfully enhanced or at least maintained antibody and T cell responses and effectively protected against SARS-CoV-2 variants in mice and NHPs. An additional heterologous booster in mice further improved and prolonged the spike-specific antibody response and conferred effective neutralizing activity against the Omicron variant. Interestingly, priming with ChAd-S and boosting with IV reduced the lung injury risk caused by T cell over activation in NHPs compared to homologous ChAd-S regimen, meanwhile maintained the flexibility of antibody regulation system to react to virus invasion by upregulating or preserving antibody levels. This study demonstrated the satisfactory compatibility of ChAd-S and IV in prime-boost vaccination in animal models.

NaOH, dolomite and NiCl2 were used as catalysts to examine their effects on co–pyrolysis with waste bicycle tires (WT) and waste engine oil (WEO). The pyrolysis behaviors with catalysts were investigated by thermogravimetric analysis. The activation energy of the catalytic main reaction stage was derived by the Kissinger–Akahira–Sunose (KAS) method under four different heating rates conditions. The calculations show that all three catalysts can reduce the activation energy of the reaction. Co–pyrolysis of WT and WEO with different catalysts was performed in a self–made lab bench at 600 °C to explore the impact on the distribution of three–phase products. The properties of gas and oil products were characterized by FTIR and Py–GC/MS (Agilent 7890B, Santa Clara, CA, USA). With the mixing of catalysts, activation energy (Eα) decreased by 15–30% in the main reaction process. NaOH and dolomite increased the yield of gas by 7% and 10%. NaOH can significantly improve the yield of CH4. The proportion of limonene in pyrolysis oil increased to 19.65% with 10% NaOH. This article provides a new method for efficiently producing limonene by mixing WT and WEO with NaOH.

Protostellar outflows are considered a signpost of star formation. These outflows can cause shocks in the molecular gas and are typically traced by the line wings of certain molecules. HNCO (4–3) has been regarded as a shock tracer because of the high abundance in shocked regions. Here we present the first imaging results of HNCO (4–3) line wings toward nine sources in a sample of 23 massive star-forming regions using the Instituto de Radioastronomía Milimétrica 30 m Telescope. We adopt the velocity range of the full width of HC _3 N (10–9) and H ^13 CO ^+ (1–0) emissions as the central emission values, beyond which the emission from HNCO (4–3) is considered to be from line wings. The spatial distributions of the red and/or blue lobes of HNCO (4–3) emission nicely associate with those lobes of HCO ^+ (1–0) in most of the sources. High-intensity ratios of HNCO (4–3) to HCO ^+ (1–0) are obtained in the line wings. The derived column density ratios of HNCO to HCO ^+ are consistent with those previously observed toward massive star-forming regions. These results provide direct evidence that HNCO could trace outflow in massive star-forming regions. This work also implies that the formation of some HNCO molecules is related to shock, either on the grain surface or within the shocked gas.

Ions (e.g., ${{\rm{H}}}_{3}^{+}$ , H _2 O ^+ ) have been used extensively to quantify the cosmic-ray ionization rate (CRIR) in diffuse sightlines. However, measurements of the CRIR in low-to-intermediate density gas environments are rare, especially when background stars are absent. In this work, we combine molecular line observations of CO, OH, CH, and HCO ^+ in the star-forming cloud IC 348, and chemical models to constrain the value of the CRIR and study the response of the distribution of chemical abundances. The cloud boundary is found to have an A _V of approximately 4 mag. From the interior to the exterior of the cloud, the observed ^13 CO line intensities drop by an order of magnitude. The calculated average abundance of ^12 CO (assuming ^12 C/ ^13 C=65) is (1.2 ± 0.9) × 10 ^−4 , which decreases by a factor of 6 from the interior to the outside regions. The average abundance of CH (3.3 ± 0.7 × 10 ^−8 ) is in good agreement with previous findings in diffuse and translucent clouds ( A _V < 5 mag). However, we did not find a decline in CH abundance in regions of high extinction ( A _V ≃ 8 mag) as previously reported in Taurus. By comparing the observed molecular abundances and chemical models, we find a decreasing trend of the CRIR as A _V increases. The inferred CRIR of ζ _cr = (4.7 ± 1.5) × 10 ^−16 s ^−1 at low A _V is consistent with ${{\rm{H}}}_{3}^{+}$ measurements toward two nearby massive stars.

The cosmic-ray ionization rate (CRIR, ζ _2 ) is one of the key parameters controlling the formation and destruction of various molecules in molecular clouds. However, the current most commonly used CRIR tracers, such as ${{\rm{H}}}_{3}^{+}$ , OH ^+ , and H _2 O ^+ , are hard to detect and require the presence of background massive stars for absorption measurements. In this work, we propose an alternative method to infer the CRIR in diffuse clouds using the abundance ratios of OH/CO and HCO ^+ /CO. We have analyzed the response of chemical abundances of CO, OH, and HCO ^+ on various environmental parameters of the interstellar medium in diffuse clouds and found that their abundances are proportional to ζ _2 . Our analytic expressions give an excellent calculation of the abundance of OH for ζ _2 ≤ 10 ^−15 s ^−1 , which are potentially useful for modeling chemistry in hydrodynamical simulations. The abundances of OH and HCO ^+ were found to monotonically decrease with increasing density, while the CO abundance shows the opposite trend. With high-sensitivity absorption transitions of both CO (1–0) and (2–1) lines from Atacama Large Millimeter/submillimeter Array, we have derived the H _2 number densities ( ${n}_{{{\rm{H}}}_{2}}$ ) toward 4 lines of sight; assuming a kinetic temperature of T _k = 50 K, we find a range of (0.14 ± 0.03–1.2 ± 0.1) × 10 ^2 cm ^−3 . By comparing the observed and modeled HCO ^+ /CO ratios, we find that ζ _2 in our diffuse gas sample is in the range of ${1.0}_{-1.0}^{+14.8}$ × 10 ${}^{-16}\mbox{--}{2.5}_{-2.4}^{+1.4}$ × 10 ^−15 s ^−1 . This is ∼2 times higher than the average value measured at higher extinction, supporting an attenuation of CRs as suggested by theoretical models.

Karst leakage is the key problem that restricts the construction of reservoir areas. In this article, the hydrogeochemical origin and hydraulic connection of the river water, pore water, fissure water, and karst water in Jinghe Dongzhuang Reservoir, which is located in a karst area, are analyzed to determine the possibility of karst leakage in the reservoir area. Piper diagram, Gibbs diagram, ion proportion coefficient, and cluster analysis were comprehensively used to systematically study the hydrogeochemical characteristics and formation mechanism of the study area. The research results show that the water in the study area is weakly alkaline, with complex hydrogeochemical types, including SO4−Na, HCO3·SO4−Na, and HCO3·SO4−Na·Mg. Affected by evaporation and concentration, Jinghe River and shallow pore water have high TDS content, and the content of Na+(including K+), Cl− and SO42− is significantly higher than that of fissure water and karst water. Fissure water and karst water are significantly weathered by rocks, and their Ca2+ and Mg2+ mainly come from carbonate rock dissolution. In the process of groundwater evolution, cation exchange occurs more or less in the three groundwater bodies, resulting in different cation contents in different water bodies. In general, Jinghe River is similar to most of the pore water, but its hydrogeochemical characteristics are obviously different from those of fissure water and karst water, so it has little hydraulic connection with fissure water and karst water, indicating that the leakage in the reservoir area is not significant.

Landscape connectivity refers to the degree of continuity between the spatially structured units of a landscape. Ecological connectivity can characterise the degree to which ecological functional areas are connected in terms of function and ecological processes. In this study, the landscape pattern index and ecosystem service values were used to evaluate the ecological functional resistance of each landscape type, taking the Three Parallel Rivers Natural World Heritage Site as an example and the habitat distribution and population size of the Yunnan snub-nosed monkey as a reference. The minimum cost distance model, combined with the barrier impact index (BEI) and ecological connectivity index (ECI), was used to determine the degree of barrier impact on the study area and the ecological connectivity of the core reserve of the heritage site in both 2000 and 2020. The resistances of the different land types and landscape heterogeneity to the ecological function of species migration between the core protected areas of the heritage site were, in descending order, those of the forest, shrubs and grass, water, unused land, cultivated land, and built-up land. In 2020, the study area had a large BEI, with areas such as built-up areas, major roads, the sides of large rivers, and arable land being significant contributors to the blockage of landscape connectivity. The overall landscape connectivity in the study area was generally low, with clear spatial differentiation and a three-column parallel distribution pattern influenced by the topography and landscape. With the adjustment of the core reserve boundaries of the heritage site, the proportion of areas with high connectivity (ECI = 4–5) increased from 11.31% in 2000 to 34.36% in 2020. This increased landscape connectivity was conducive to the migration and reproduction of large terrestrial animals, such as the Yunnan snub-nosed monkey, with increasing numbers of populations and individuals. This study provides theoretical and methodological insights into the delineation and conservation of natural heritage sites and landscape connectivity.

A rapid shrinkage of Daihai Lake was found in recent decades. The present study analyzed the characteristics of Daihai Lake shrinkage and quantified the contribution of climate and human activities. The results of Mann-Kendall- Sneyers test and moving t-test showed that there was an obvious mutation point of lake level in 2006 and the descending speed of Daihai Lake level post-2006 (-0.46m/a) was 3.22 times that of pre-2006 (-0.14m/a). The centroid of Daihai Lake moved 1365.18 m from southwest to northeast during 1989 ~ 2018 with an average speed of 47.08 m/a. The results of Mann-Kendall trend test revealed that the annual evaporation showed a significant downward trend with a rate of approximately -5.33 mm/a, while no significant trend was found in precipitation. Daihai lake water level showed a very weak relationship with evaporation (r = 0.078, p < 0.01) and precipitation (p>0.05) respectively. Daihai Lake was influenced by human activities mainly from land use/ land cover, building reservoirs, pumping groundwater and directly consuming Daihai Lake water by Daihai power plant (DHPP). It was thought-provoking that DHPP began to consume Daihai lake water in 2006, which was consistent with abrupt change of Daihai lake level. The proportion of human impact was fluctuating upward. Human factors were the main factor of lake water reduction in last 10 years and the 5-year average contribution of human activities to Daihai Lake shrinkage was more than 61.99%. More attention and economic support should be given to prevent the continuous shrinkage of Daihai Lake.

Herbicide resistance to chemical herbicide is a global issue that presents an ongoing threat to grain production. Though it has been frequently implicated that the production of detoxification enzymes increased in resistance development, the mechanisms for overexpression of these genes employed by herbicide-resistant weeds remain complicated. In this study, a mesosulfuron-methyl resistant Beckmannia syzigachne population (R) was found to be cross-resistant to another herbicide pyriminobac-methyl. No known target-site mutations were detected in the R population. In contrast, the decreased uptake and enhanced metabolic rates of mesosulfuron-methyl were detected in the R than the susceptible (S) population. Two candidate ATP-binding cassette (ABC) transporter genes (ABCB25 and ABCC14) that were constitutively up-regulated in the R population were identified by RNA-sequencing and validated by RT-qPCR. Alteration of antioxidant enzyme activities and gene expressions implied that mesosulfuron-methyl-induced antioxidant defenses provoked reactive oxygen species (ROS) burst. ROS scavenger assay showed that ROS induces ABCB25 and ABCC14 expression. This study reported for the first time that ABC transporters mediated non-target-site resistance contributes to mesosulfuron-methyl resistance in a B. syzigachne population, and implicated that ROS burst might be involved in the overexpression of ABC transporter genes in weeds.

Malus hupehensis (Pamp.) Rehd (MH) is a dual-purpose plant used as medicine and food, which possesses various biological activities including hepatoprotective effect. In this study, the hepatoprotective activity of extracts of MH (MHE) against RFP + INH-induced liver injury was investigated. Animals in model group were given saline and INH + RFP. Animals in MHE groups were given MHE and INH + RFP. Animals in normal control group were given equal volume of saline. Subsequently, blood and liver samples were collected and assessed. MHE significantly alleviated the liver injury, as evidenced by decreased activities of the ALT, AST, TBIL, MDA and GSH-ST, and increased levels of SOD and GSH. MHE also effectively reduced the pathological tissue damage. Moreover, the CYP450 levels, and expression of CYP3A4, CYP2C9 and NADPH4 protein were inhibited by MHE. MHE exerts a protective effect against RFP + INH-induced liver injury in mice, and could be developed as a liver protectant for treatment of DILI.

Alopecurus japonicus has been a serious weed across China and developed resistance to the acetyl-CoA carboxylase (ACCase)-inhibiting herbicide. The A. japonicus ACCase gene mutations accounting for target-site resistance (TSR) have been clarified, while non-target-site resistance (NTSR) is not distinct. Here, dose–response testing indicated that an A. japonicus population (R) was resistant to four ACCase-inhibiting herbicides fenoxaprop-P-ethyl, sethoxydim, clethodim, and pinoxaden. Compared with herbicide-susceptible A. japonicus (S), no known resistant mutations for TSR in ACCase were identified in the R population using sequencing. To investigate the NTSR mechanism, increased metabolism of fenoxaprop-P-ethyl was detected in the R population using high-performance liquid chromatography (HPLC) analysis. Notably, resistance cannot be reversed by P450 and GST inhibitors. RNA-seq was performed to further explore the resistance mechanisms, and eight candidate contigs (four glycosyl transferases (GT) and four ATP-binding cassette (ABC) transporters) were chosen and their expression patterns were validated using RT-qPCR. Three GT and three ABC transporter contigs were constitutively upregulated in the R population. In short, six contigs expressed highly in the R population causing enhanced fenoxaprop-P-ethyl metabolism appear to be involved in fenoxaprop-P-ethyl resistance.

Real-time matching of millimeter wave (mmWave) narrow beams is a great challenge in dynamic vehicle to infrastructure (V2I) environments due to the mobility of vehicles. In this paper, a novel beam search strategy based on spectrum-environment awareness is proposed. Combining the technique of label iterative optimization with three-dimensional (3D) grid encoding, the strategy treats the optimal beam pair indexes (BPIs) as labels and encodes the environments as features. Three-dimensional grid encoding is a symmetry-based environmental coding technology. A new Convolutional Neural Network (CNN) model is also constructed, which is trained by the features. The situational beam search of actual vehicles is performed under the trained CNN model. As a result, real-time mmWave narrow beam matching can be achieved. Simulation results demonstrate that the proposed strategy can effectively reduce the beam search overhead and improve the efficiency while guaranteeing the matching accuracy.

Urbanization improves the living environment of billions of people but negatively impacts the local carbon cycle as it encroaches on ecosystems. We analyzed this dual effect of urbanization by quantifying linkages between several indicators -- urbanization-induced terrestrial carbon storage loss (uTCSL), per capita urban area (PCUA), and urban land use efficiency (LCRPGR) -- within the Hohhot-Baotou-Ordos (HBO) region in 1990-2100. Results show that the growth of 4.45 million urban residents and 420.70 km² urban area led to a uTCSL of 815.74 GgC in the HBO region in 1990-2020. The uTCSL contributed to improving the urban crowding indicated by PCUA but was subject to LCRPGR. The PCUA decreased by -33.75 m²/person under high land use efficiency (LCRPGR<1) in 1990-2010. Although the PCUA increased by 23.59 m²/person from 2010 to 2020, it resulted in a 2.08-fold increase in uTCSL compared to 1990-2010, with each additional 1 m² increase in PCUA being associated with carbon storage loss of 24.50 GgC. Our projections indicate that regulating LCRPGR twenty years before the urban population reaches its peak could benefit both uTCSL and PCUA. This study provides valuable insights for urban planning aligned with the UN 2030 Agenda, especially for arid and semi-arid cities. [ABSTRACT FROM AUTHOR]