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1.
Front Cardiovasc Med ; 11: 1381391, 2024.
Article in English | MEDLINE | ID: mdl-38737715

ABSTRACT

Background: The incidence of deep vein thrombosis (DVT) in the lower extremities is increasing in the younger population. However, there are fewer reported comparisons in the literature for lower extremity DVT. Methods: Patients aged <40 years admitted with lower-extremity DVT between January 2018 and December 2023 were retrospectively analyzed and followed up for 1 year. Results: A total of 61 patients were included in the study and divided into two groups: 33 patients over 30 years of age (middle-aged group) and 28 patients under 30 years of age (young group). A significant gender difference was observed, with a higher proportion of males in the young group compared to the middle-aged group (P < 0.001). Five patients in the young group were treated with anticoagulation alone, whereas all patients in the middle-aged group underwent endovascular therapy. A higher prevalence of inferior vena cava thrombosis in the young group compared to the middle-aged group (60.71% vs. 33.3%, P = 0.032). The proportion of iliac vein stenosis was significantly higher in the middle-aged groups than in the young group (P = 0.002). There was no statistically significant difference in venous function scores (Villalta and rVCSS) between the two groups during both the preoperative period and the postoperative follow-up (P > 0.05). The incidence of lower-extremity DVT post-thrombotic syndrome and thrombus recurrence was higher in the young group than in the middle-aged group at 1 year postoperatively (PTS: 78.57% vs. 33.3%, P < 0.001, and thrombus recurrence: 28.57% vs. 9.09%, P < 0.05). Univariate and multivariate analyses revealed that inferior vena cava thrombosis was an independent risk factor for severe DVT post-thrombotic syndrome and recurrent DVT (P < 0.05), whereas gender was an independent risk factor for recurrent DVT (P < 0.05). Conclusions: This study suggests differences in the clinical characteristics and prognosis of lower-extremity DVT.

2.
J Mech Behav Biomed Mater ; 141: 105764, 2023 05.
Article in English | MEDLINE | ID: mdl-36965216

ABSTRACT

Temporomandibular joint disorder (TMD) often coincides with malocclusion, and in some cases TMDs are reported after orthodontic treatment. Intermaxillary elastics (also known as orthodontic elastics, OE) are a common way to apply force during orthodontic treatment, and they might cause mechanical effects on the temporomandibular joint (TMJ), thereby lead to joint remodeling. It is still a controversial topic whether the adapted remodeling of the TMJ or of the alveolar bone is the main cause for the alteration of occlusion after treatment with OEs. It was the aim of this study to analyze whether variations of OEs would develop harmful effects on the healthy TMJ. A TMJ model with a masticatory system based on Hill-type muscle actuators was established. Mouth opening and closure with and without OEs were simulated, and maximum principal stresses in the disc and condylar cartilage as well as the displacement of the mandible were analyzed. We found no considerably difference in the mandibular movement without and with symmetrical OEs during mouth opening and closing. At full mouth opening, stresses in the disc and condylar cartilage of some models with OEs were much smaller than without OEs, but we did not find consistency in the results from the left and right sides of the same model (e.g. the lowest compressive stress on the left side of disc from the model with Class II OEs is much smaller than without OEs, -17.3 MPa compared with -28.2, while on the right side, there was no obvious difference). Hence, we could not conclude that OEs would develop deleterious effects on the healthy TMJ.


Subject(s)
Temporomandibular Joint Disc , Temporomandibular Joint Disorders , Humans , Temporomandibular Joint/physiology , Mandible , Movement , Mandibular Condyle
3.
Vaccines (Basel) ; 10(11)2022 Nov 12.
Article in English | MEDLINE | ID: mdl-36423012

ABSTRACT

Conjugate-vaccine immunogens require three components: a carrier protein, an antigen, and a crosslinker, capable of coupling antigen to carrier protein, while preserving both T-cell responses from carrier protein and B-cell responses from antigen. We previously showed that the N-terminal eight residues of the HIV-1 fusion peptide (FP8) as an antigen could prime for broad cross-clade neutralizing responses, that recombinant heavy chain of tetanus toxin (rTTHC) as a carrier protein provided optimal responses, and that choice of crosslinker could impact both antigenicity and immunogenicity. Here, we delve more deeply into the impact of varying the linker between FP8 and rTTHC. In specific, we assessed the physical properties, the antigenicity, and the immunogenicity of conjugates for crosslinkers ranging in spacer-arm length from 1.5 to 95.2 Å, with varying hydrophobicity and crosslinking-functional groups. Conjugates coupled with different degrees of multimerization and peptide-to-rTTHC stoichiometry, but all were well recognized by HIV-fusion-peptide-directed antibodies VRC34.01, VRC34.05, PGT151, and ACS202 except for the conjugate with the longest linker (24-PEGylated SMCC; SM(PEG)24), which had lower affinity for ACS202, as did the conjugate with the shortest linker (succinimidyl iodoacetate; SIA), which also had the lowest peptide-to-rTTHC stoichiometry. Murine immunizations testing seven FP8-rTTHC conjugates elicited fusion-peptide-directed antibody responses, with SIA- and SM(PEG)24-linked conjugates eliciting lower responses than the other five conjugates. After boosting with prefusion-closed envelope trimers from strains BG505 clade A and consensus clade C, trimer-directed antibody-binding responses were lower for the SIA-linked conjugate; elicited neutralizing responses were similar, however, though statistically lower for the SM(PEG)24-linked conjugate, when tested against a strain especially sensitive to fusion-peptide-directed responses. Overall, correlation analyses revealed the immunogenicity of FP8-rTTHC conjugates to be negatively impacted by hydrophilicity and extremes of length or low peptide-carrier stoichiometry, but robust to other linker parameters, with several commonly used crosslinkers yielding statistically indistinguishable serological results.

4.
Fish Shellfish Immunol ; 124: 47-55, 2022 May.
Article in English | MEDLINE | ID: mdl-35367379

ABSTRACT

A previous study confirmed that spotted sea bass (Lateolabrax maculatus), an economically important cultured species in East Asia, is a new host of Aeromonas veronii, which can cause acute death in these fish, but there is little in-depth understanding of this disease. In the present study, the virulence of 10 isolates of A. veronii derived from spotted sea bass was determined. It was found that the 18BJ181 isolate was a virulent strain and led to the fastest death of spotted sea bass. Death was determined to be within in 2-12 h, and resulted in abdominal effusion and varying degrees of hemorrhage in internal organs. Bacterial colonization analysis showed that the bacterial load in the spleen was highest, and was up to 3.1 × 105 cfu g-1. In addition, the bacteria proliferated massively in the blood and reached 2.4 × 107 cfu mL-1 at 12 h after 18BJ181 strain infection, which was also a typical feature of acute septicemia. Histopathology of the spleen revealed edema in interstitial tissue, degeneration, and necrosis in lymphoid tissue, and hemorrhage in the capillary network. Transcriptome analysis of the spleen showed that the expression level of HSP70, CCL19, and IL-1ß was extremely significantly up-regulated at 8 h after infection (P < 0.01), and the expression of these genes was normal at 24 h. These results revealed that A. veronii infection could rapidly activate the chemokine signal pathway and stimulate the acute inflammatory response in the host. The bacterial colonization, pathological features, and gene expression patterns in immune pathways will help us to better understand acute septicemia in spotted sea bass caused by A. veronii.


Subject(s)
Bass , Fish Diseases , Sepsis , Aeromonas veronii/genetics , Animals , Gene Expression Profiling , Immunity , Sepsis/veterinary
5.
Foods ; 11(6)2022 Mar 14.
Article in English | MEDLINE | ID: mdl-35327257

ABSTRACT

Grain drying is a complex heat and mass transfer process, which has the characteristics of a significant delay, multidisturbance, nonlinearity, strong coupling, and parameter uncertainty. Artificial intelligence (AI) control technology is suitable for solving such complex control problems. In this paper, the mechanism and data dual-drive with equivalent accumulated temperature (EAT) mutual-window AI-control method for continuous grain drying were proposed, and a control system was established. The experimental verification was carried out on the test platform of continuous grain drying. The results show that the method has the ability of implicit prediction, high accuracy, strong stability and self-adaptive ability, and the maximum control deviation of moisture at the outlet of the dryer is -0.58-0.3%.

6.
Can J Microbiol ; 68(3): 215-226, 2022 Mar.
Article in English | MEDLINE | ID: mdl-34855458

ABSTRACT

Vibrio harveyi can cause infections and diseases in a variety of marine vertebrates and invertebrates, which are harmful to the aquaculture industry. The LuxS quorum-sensing system regulates the expression of virulence factors in a wide variety of pathogenic bacteria. In this study, an in-frame deletion of the luxS gene was constructed to reveal the role of LuxS in the physiology and virulence of V. harveyi. Statistical analysis showed no significant differences in the growth ability, biofilm formation, antibiotic susceptibility, virulence by intraperitoneal injection, and ability of V. harveyi to colonize the spleen and liver of the pearl gentian grouper between the wild-type (WT) and luxS mutant. However, deletion of luxS decreased the secretion of extracellular protease, while increasing swimming and swarming abilities. Simultaneously, a luxS-deleted mutant showed overproduction of lateral flagella, and an intact luxS complemented this defect. Since motility is flagella dependent, 16 V. harveyi flagella biogenesis related genes were selected for further analysis. Based on quantitative real-time reverse transcription-PCR (qRT-PCR), the expression levels of these genes, including the polar flagella genes flaB, flhA, flhF, flhB, flhF, fliS, and flrA and the lateral flagella genes flgA, flgB, fliE, fliF, lafA, lafK, and motY, were significantly upregulated in the ΔluxS: pMMB207 (ΔluxS+) strain as compared with the V. harveyi 345: pMMB207 (WT+) and C-ΔluxS strains during the early, mid-exponential, and stationary growth phases. Our results indicate that LuxS plays an important role in controlling motility, flagella biogenesis, and extracellular protease secretion in V. harveyi.


Subject(s)
Peptide Hydrolases , Vibrio , Animals , Bacterial Proteins/metabolism , Carbon-Sulfur Lyases/metabolism , Gene Expression Regulation, Bacterial , Peptide Hydrolases/genetics , Peptide Hydrolases/metabolism , Vibrio/genetics
7.
Front Genet ; 13: 1100016, 2022.
Article in English | MEDLINE | ID: mdl-36685871

ABSTRACT

Over the past decade, with the development of high-throughput single-cell sequencing technology, single-cell omics has been emerged as a powerful tool to understand the molecular basis of cellular mechanisms and refine our knowledge of diverse cell states. They can reveal the heterogeneity at different genetic layers and elucidate their associations by multiple omics analysis, providing a more comprehensive genetic map of biological regulatory networks. In the post-GWAS era, the molecular biological mechanisms influencing human diseases will be further elucidated by single-cell omics. This review mainly summarizes the development and trend of single-cell omics. This involves single-cell omics technologies, single-cell multi-omics technologies, multiple omics data integration methods, applications in various human organs and diseases, classic laboratory cell lines, and animal disease models. The review will reveal some perspectives for elucidating human diseases and constructing animal models.

8.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-923368

ABSTRACT

Objective @# To investigate the efficacy of a holistic approach for postoperative pain management in children receiving dental treatment under general anesthesia in day-surgery operating room. @*Methods@#A total of 120 children, aged 3-7 years, of American Society of Anesthesiologists physical status Ⅰ or Ⅱ, with ≥ 10 treated teeth, receiving comprehensive dental treatment under general anesthesia from January 2020 to August 2020 were enrolled in this trial and randomly allocated into the holistic approach group (group H, n=60) and including preemptive analgesia, instructions to parents for pain management and web-based assessment system (assessment pain by scanning the quick response code 4, 6, and 24 hours postoperatively) and the control group (group C, n=60) only scanning the quick response code. Pain, face, legs, activity, cry and consolability (FLACC) scale was used to assess the level of pain 2 h postoperatively and the parents postoperative pain measure (PPPM) was used to assess the level of pain 4, 6, and 24 h postoperatively in two groups.@* Results@# The FLACC scores of group H 2 h postoperatively were significantly lower than group C (P <0.05). The incidences of significant pain (PPPM scores ≥ 6) 4, 6 and 24 h postoperatively in group H were lower than group C (P <0.05). Altogether, 91.7% of parents in group H and 71.6% in group C assessed the level of pain of children over time. The compliance rate of parents in group H was significantly higher than group C (P <0.05).@*Conclusion @# The holistic approach had a positive effect on reducing postoperative pain for children receiving dental treatment under general anesthesia in the day-surgery operating room.

9.
Sci Rep ; 11(1): 18663, 2021 09 20.
Article in English | MEDLINE | ID: mdl-34545147

ABSTRACT

Drought resistance of psammophyte determines survival and growth, but their responses to drought are not well understood. We conducted a pot experiment to study how physiological characteristics respond to drought and rehydration. We found that watering to 60-65% of field capacity (the control) provided more water than was required by Agriophyllum squarrosum and its leaves became yellow and slightly wilted. The total chlorophyll content and Fm (maximum fluorescence after dark adaptation) in control were lower than in the drought treatment, and both decreased after rehydration. With increasing drought duration and intensity, the relative water content (RWC), chlorophyll content, Fm, and the quantum efficiency of photosystem II (Fv/Fm) of Setaria viridis decreased, but malondialdehyde and membrane permeability increased. During the late drought, the activities of three antioxidant enzymes in A. squarrosum increased to prevent membrane lipid peroxidation; for S. viridis, only peroxidase and superoxide dismutase activities increased. After rehydration, RWC of both species increased, but Fv/Fm of A. squarrosum and Fm of S. viridis did not recover under severe drought. Our research illustrated that A. squarrosum is better adapted to arid environment than S. viridis, but the high soil moisture content is not conducive to normal growth of A. squarrosum.


Subject(s)
Amaranthaceae/metabolism , Setaria Plant/metabolism , Stress, Physiological/physiology , Adaptation, Physiological/physiology , Amaranthaceae/growth & development , Amaranthaceae/physiology , Antioxidants/metabolism , Chlorophyll , Conservation of Natural Resources/methods , Droughts , Environmental Restoration and Remediation/methods , Mongolia , Peroxiredoxins/metabolism , Photosynthesis/physiology , Plant Leaves/metabolism , Setaria Plant/growth & development , Setaria Plant/physiology , Soil/chemistry , Water/analysis
10.
Environ Sci Pollut Res Int ; 28(48): 68174-68187, 2021 Dec.
Article in English | MEDLINE | ID: mdl-34264496

ABSTRACT

As the global ecosystem has been severely disturbed by an increasing number of human activities at different scales, remote sensing technology, as an effective quantitative measure of environmental quality, has been widely used. The remote sensing ecological index (RSEI) is one of the most popular and comprehensive ecological quality assessment indices based on the remote sensing data. However, the RSEI model exhibits that the ecological environment under natural conditions is not limited by the spatial scales. In addition, the model has major shortcomings in index selection and eigenvector, which greatly limit the application of RSEI. In this paper, the RSEI model is improved and a remote sensing ecological index optimized by the regional scale (RO-RSEI) is proposed. The result of the study, conducted in Shuangyang District, Changchun City, Jilin Province, shows that the RO-RSEI model has regional ecological significance after the introduction of the scale theory of landscape ecology; the index is preferred to solve problems like the RSEI model applied mechanization and baseless index selection. Meanwhile, due to the optimization of the eigenvector contribution of the optimal index, it solves the problems like non-unique model calculation result caused by principal component analysis or even antipodal calculation result. Compared with the RSEI model, the monitoring result of RO-RSEI model can better reflect the regional ecological changes. The improved model offers the possibility of monitoring ecological environment quality with remote sensing big data and provides a scientific basis for future scholars' batch computing.


Subject(s)
Ecosystem , Environmental Monitoring/methods , Remote Sensing Technology , China , Cities , Environment
11.
Vaccine ; 39(25): 3379-3387, 2021 06 08.
Article in English | MEDLINE | ID: mdl-34020817

ABSTRACT

Metastable glycosylated immunogens present challenges for GMP manufacturing. The HIV-1 envelope (Env) glycoprotein trimer is covered by N-linked glycan comprising half its mass and requires both trimer assembly and subunit cleavage to fold into a prefusion-closed conformation. This conformation, the vaccine-desired antigenic state, is both metastable to structural rearrangement and labile to subunit dissociation. Prior reported GMP manufacturing for a soluble trimer stabilized in a near-native state by disulfide (SOS) and Ile-to-Pro (IP) mutations has employed affinity methods based on antibody 2G12, which recognizes only ~30% of circulating HIV strains. Here, we develop a scalable manufacturing process based on commercially available, non-affinity resins, and we apply the process to current GMP (cGMP) production of trimers from clades A and C, which have been found to boost cross-clade neutralizing responses in vaccine-test species. The clade A trimer, which we named "BG505 DS-SOSIP.664", contained an engineered disulfide (201C-433C; DS) within gp120, which further stabilized this trimer in a prefusion-closed conformation resistant to CD4-induced triggering. BG505 DS-SOSIP.664 was expressed in a CHO-DG44 stable cell line and purified with initial and final tangential flow filtration steps, three commercially available resin-based chromatography steps, and two orthogonal viral clearance steps. The non-affinity purification enabled efficient scale-up, with a 250 L-scale cGMP run yielding 9.6 g of purified BG505 DS-SOSIP.664. Antigenic analysis indicated retention of a prefusion-closed conformation, including recognition by apex-directed and fusion peptide-directed antibodies. The developed manufacturing process was suitable for 50 L-scale production of a second prefusion-stabilized Env trimer vaccine candidate, ConC-FP8v2 RnS-3mut-2G-SOSIP.664, yielding 7.8 g of this consensus clade C trimer. The successful process development and purification scale-up of HIV-1 Env trimers from different clades by using commercially available materials provide experimental demonstration for cGMP manufacturing of trimeric HIV-Env vaccine immunogens, in an antigenically desired conformation, without the use of costly affinity resins.


Subject(s)
AIDS Vaccines , HIV-1 , Antibodies, Neutralizing , HIV Antibodies , HIV Antigens , HIV-1/genetics , Protein Multimerization , env Gene Products, Human Immunodeficiency Virus/genetics
12.
Microb Pathog ; 157: 104965, 2021 Aug.
Article in English | MEDLINE | ID: mdl-34015493

ABSTRACT

Vibrio harveyi is a Gram-negative bacterium that occurs widely in the ocean and a kind of pathogenic bacteria associated with vibriosis in grouper. We investigated whether the VscCD protein of the type Ⅲ secretion system (T3SS) was important for pathogenicity of V. harveyi. Mutations to the vscC and vscD genes (ΔvscCD) and complementation of the ΔvscCD mutant (C-ΔvscCD) were created. Moreover, the biological characteristics of the wild-type (WT) and mutant strains of V. harveyi 345 were compared. The results showed that deletion of the vscCD genes had no effect on bacterial growth, swimming/swarming ability, secretion of extracellular protease, or biofilm formation. However, as compared with the V. harveyi 345: pMMB207 (WT+) and complementary (C-ΔvscCD) strains, the ΔvscCD: pMMB207 (ΔvscCD+) mutant displayed decreased resistance to acid stress, H2O2, and antibiotics. In addition, infection of the pearl gentian grouper (♀Epinephelus fuscoguttatus × â™‚Epinephelus lanceolatu) showed that as compared with the WT+ and C-ΔvscCD strains, the ΔvscCD+ strain significantly reduced cumulative mortality of the host. The colonization ability of the ΔvscCD+ mutant in the spleen and liver tissues of the pearl gentian grouper was significantly lower than that of the WT+ and C-ΔvscCD strains. In the early stage of infection with the ΔvscCD+ strain, the expression levels of IL-1ß, IL-16, TLR3, TNF-α, MHC-Iα, and CD8α were up-regulated to varying degrees. As compared with the WT+ and C-ΔvscCD strains, luxR expression was significantly up-regulated in the ΔvscCD+ strain, while the expression of vcrH and vp1668 was significantly down-regulated. As an important component of the T3SS, VscCD seemed to play a significant role in the pathogenesis of V. harveyi.


Subject(s)
Fish Diseases , Vibrio Infections , Vibrio , Animals , Hydrogen Peroxide , Type III Secretion Systems/genetics , Vibrio/genetics , Vibrio Infections/veterinary
13.
Foods ; 11(1)2021 Dec 31.
Article in English | MEDLINE | ID: mdl-35010228

ABSTRACT

In our study, we developed a system to reduce both energy consumption and pollutant discharge during the drying process. We present a new technology, a stationary bed grain-drying test device based on the internal circulation of the drying medium (ICODM). A rice-drying experiment was carried out inside of it, and the influences of air temperature (AT) and air velocity (AV) on the energy and exergy efficiencies (EEE) as well as the improvement potential rate (IPR) and the sustainability index (SI) of the rice-drying process were studied. The following conclusions were obtained: when the rice was dried at a temperature of below 55 °C and an AV across the grain layer of 0.5 m/s, the average EEE during the drying process was 48.27-72.17% and 40.27-71.07%, respectively, demonstrating an increasing trend as the drying medium temperature increased. When the rice was dried using an AV across the grain layer in the range of 0.33-0.5 m/s and a temperature of 40 °C, the two values were 39.79-73.9% and 49.66-71.04%, respectively, demonstrating a decreasing trend as the drying medium flow velocity increased. IPR and SI were 4.1-8.5 J/s and 1.9-2.7, respectively, at a drying temperature of 30-55 °C and an AV of 0.33-0.5 m/s. These conclusions can provide helpful guidance for the optimization and control of the rice-drying process in terms of saving energy.

14.
Front Immunol ; 11: 607754, 2020.
Article in English | MEDLINE | ID: mdl-33324424

ABSTRACT

Vibrio harveyi causes vibriosis in nearly 70% of grouper (Epinephelus sp.), seriously limiting grouper culture. As well as directly inhibiting pathogens, the gut microbiota plays critical roles in immune homeostasis and provides essential health benefits to its host. However, there is still little information about the variations in the immune response to V. harveyi infection and the gut microbiota of grouper. To understand the virulence mechanism of V. harveyi in the pearl gentian grouper, we investigated the variations in the pathological changes, immune responses, and gut bacterial communities of pearl gentian grouper after exposure to differently virulent V. harveyi strains. Obvious histopathological changes were detected in heart, kidney, and liver. In particular, nodules appeared and huge numbers of V. harveyi cells colonized the liver at 12 h postinfection (hpi) with highly virulent V. harveyi. Although no V. harveyi was detected in the gut, the infection simultaneously induced a gut-liver immune response. In particular, the expression of 8 genes associated with cellular immune processes, including genes encoding inflammatory cytokines and receptors, and pattern recognition proteins, was markedly induced by V. harveyi infection, especially with the highly virulent V. harveyi strain. V. harveyi infection also induced significant changes in gut bacterial community, in which Vibrio and Photobacterium increased but Bradyrhizobium, Lactobacillus, Blautia, and Faecalibaculum decreased in the group infected with the highly virulent strain, with accounting for 82.01% dissimilarity. Correspondingly, four bacterial functions related to bacterial pathogenesis were increased by infection with highly virulent V. harveyi, whereas functions involving metabolism and genetic information processing were reduced. These findings indicate that V. harveyi colonizes the liver and induces a gut-liver immune response that substantially disrupts the composition of and interspecies interactions in the bacterial community in fish gut, thereby altering the gut-microbiota-mediated functions and inducing fish death.


Subject(s)
Fish Diseases/microbiology , Fishes/microbiology , Gastrointestinal Microbiome , Gastrointestinal Tract/microbiology , Liver/microbiology , Vibrio Infections/veterinary , Vibrio/pathogenicity , Animals , Dysbiosis , Female , Fish Diseases/genetics , Fish Diseases/immunology , Fish Diseases/metabolism , Fishes/genetics , Fishes/immunology , Fishes/metabolism , Gastrointestinal Tract/immunology , Gastrointestinal Tract/metabolism , Gene Expression Regulation, Bacterial , Host-Pathogen Interactions , Liver/immunology , Liver/metabolism , Male , Vibrio/genetics , Vibrio/immunology , Vibrio Infections/immunology , Vibrio Infections/metabolism , Vibrio Infections/microbiology , Virulence
15.
Vaccine ; 38(29): 4507-4511, 2020 06 15.
Article in English | MEDLINE | ID: mdl-32448620

ABSTRACT

An N-terminal peptide of the HIV-1 fusion peptide (FP) with eight amino acid residues (FP8) was conjugated to a recombinant Tetanus Toxoid Heavy Chain Fragment C (rTTHc) as a carrier protein to help boosting immunogenicity against HIV-1. In this rapid communication, a unique algorithm to determine FP-rTTHc conjugation ratio was developed based off the amino acid analysis. Five well recovered amino acids (present in both FP and rTTHc) were used to calculate the conjugation ratio, while proline (present only in rTTHc) was identified and utilized as the intrinsic internal standard for normalization. With this calculation, the assay variability was minimized (<20%), especially for conjugates with moderate to low conjugation ratios as being compared to previously reported methods. The approach offers a reliable tool to determine the efficiency of the conjugation reactions for in-process monitoring and for final conjugate product characterization.


Subject(s)
Amino Acids , Carrier Proteins , Algorithms , Peptides , Reference Standards
16.
Article in English | MEDLINE | ID: mdl-32224438

ABSTRACT

A fusion peptide mimicking a part of the sequence of HIV-1 envelope glycoprotein with an additional cysteine at its C-terminus (FP8: AVGIGAVFC) was conjugated to a carrier protein through a linker for development of an HIV-1 vaccine. Since this fusion peptide is very hydrophobic with poor solubility and can self-dimerize via a disulfide bond, co-existence of monomeric and dimeric forms presented a major challenge for residual unconjugated FP8 quantification. A reversed-phase liquid chromatography (RPLC) with UV detection was developed to monitor residual FP8 using an experimental correction factor of 0.85 for UV peak area measurement between FP8 dimer and monomer. Therefore, both forms of unconjugated residual FP8 can be measured based on a single FP8 monomer reference curve. Overall, this study demonstrated that the current purification process can remove free residual FP8 to a low level, <20 µg/mL, which showed negligible impact (<10%) for the conjugated FP8 ratio measurement using another method, amino acid analysis.


Subject(s)
Peptides/analysis , Recombinant Fusion Proteins/analysis , Amino Acid Sequence , Chromatography, High Pressure Liquid , Chromatography, Reverse-Phase , Cysteine/chemistry , Disulfides/chemistry , Humans , Hydrophobic and Hydrophilic Interactions , Limit of Detection , Protein Conformation , Protein Multimerization
17.
Sci Rep ; 10(1): 3032, 2020 02 20.
Article in English | MEDLINE | ID: mdl-32080235

ABSTRACT

The vaccine elicitation of broadly neutralizing antibodies against HIV-1 is a long-sought goal. We previously reported the amino-terminal eight residues of the HIV-1-fusion peptide (FP8) - when conjugated to the carrier protein, keyhole limpet hemocyanin (KLH) - to be capable of inducing broadly neutralizing responses against HIV-1 in animal models. However, KLH is a multi-subunit particle derived from a natural source, and its manufacture as a clinical product remains a challenge. Here we report the preclinical development of recombinant tetanus toxoid heavy chain fragment (rTTHC) linked to FP8 (FP8-rTTHC) as a suitable FP-conjugate vaccine immunogen. We assessed 16 conjugates, made by coupling the 4 most prevalent FP8 sequences with 4 carrier proteins: the aforementioned KLH and rTTHC; the H. influenzae protein D (HiD); and the cross-reactive material from diphtheria toxin (CRM197). While each of the 16 FP8-carrier conjugates could elicit HIV-1-neutralizing responses, rTTHC conjugates induced higher FP-directed responses overall. A Sulfo-SIAB linker yielded superior results over an SM(PEG)2 linker but combinations of carriers, conjugation ratio of peptide to carrier, or choice of adjuvant (Adjuplex or Alum) did not significantly impact elicited FP-directed neutralizing responses in mice. Overall, SIAB-linked FP8-rTTHC appears to be a promising vaccine candidate for advancing to clinical assessment.


Subject(s)
AIDS Vaccines/immunology , HIV-1/immunology , Peptides/immunology , Recombinant Fusion Proteins/immunology , Adjuvants, Immunologic , Amino Acid Sequence , Animals , Cross Reactions/immunology , Female , Immunization , Mice, Inbred BALB C , Mice, Inbred C57BL , Neutralization Tests , Peptides/chemistry
18.
Plants (Basel) ; 8(7)2019 Jul 23.
Article in English | MEDLINE | ID: mdl-31340533

ABSTRACT

The availability of water is the critical factor driving plant growth, physiological responses, population and community succession in arid and semiarid regions, thus a precipitation addition-reduction platform with five experimental treatments, was established to explore the growth and physiology of two psammophytes (also known as psammophiles) to precipitation manipulation in Horqin Sandy Land. Changes in coverage and density were measured, and antioxidant enzymes and osmoregulatory substances in both of the studied species were determined. Investigation results showed that the average vegetation coverage increased with an increasing precipitation, and reached a maximum in July. Under the -60% precipitation treatment, Tribulus terrestris accounted for a large proportion of the area, but Bassia dasyphylla was the dominant species in the +60% treatment. T. terrestris was found to have higher a drought stress resistance than B. dasyphylla. From days 4 to 7 after rainfall, B. dasyphylla under precipitation reduction showed obvious water stress. The malondialdehyde (MDA) content of B. dasyphylla was higher than that of T. terrestris, but that of B. dasyphylla had the lower relative water content (RWC). The MDA content in the precipitation reduction treatments of the two studied species was higher than that in the precipitation addition treatments from days 4 to 10. Peroxidase (POD) and superoxide dismutase (SOD) activity and the soluble proteins and free proline content of T. terrestris were higher than those of B. dasyphylla. The free proline content of T. terrestris and B. dasyphylla increased with increasing drought stress. Our data illustrated that T. terrestris had a higher drought stress resistance than B. dasyphylla, which was correlated with the augmentation of some antioxidant enzymes and osmoregulatory substance. The adaptive mechanism provides solid physiological support for an understanding of psammophyte adaptation to drought stress, and of community succession or species manipulation for desertified land restoration.

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