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1.
J Virol ; 98(2): e0165023, 2024 Feb 20.
Artículo en Inglés | MEDLINE | ID: mdl-38271227

RESUMEN

Vaccination is the most effective method to protect humans and animals from diseases. Anti-idiotype vaccines are safer due to their absence of pathogens. However, the commercial production of traditional anti-idiotype vaccines using monoclonal and polyclonal antibodies (mAb and pAb) is complex and has a high failure rate. The present study designed a novel, simple, low-cost strategy for developing anti-idiotype vaccines with nanobody technology. We used porcine circovirus type 2 (PCV2) as a viral model, which can result in serious economic loss in the pig industry. The neutralizing mAb-1E7 (Ab1) against PCV2 capsid protein (PCV2-Cap) was immunized in the camel. And 12 nanobodies against mAb-1E7 were screened. Among them, Nb61 (Ab2) targeted the idiotype epitope of mAb-1E7 and blocked mAb-1E7's binding to PCV2-Cap. Additionally, a high-dose Nb61 vaccination can also protect mice and pigs from PCV2 infection. Epitope mapping showed that mAb-1E7 recognized the 75NINDFL80 of PCV2-Cap and 101NYNDFLG107 of Nb61. Subsequently, the mAb-3G4 (Ab3) against Nb61 was produced and can neutralize PCV2 infection in the PK-15 cells. Structure analysis showed that the amino acids of mAb-1E7 and mAb-3G4 respective binding to PCV2-Cap and Nb61 were also similar on the amino acids sequences and spatial conformation. Collectively, our study first provided a strategy for producing nanobody-based anti-idiotype vaccines and identified that anti-idiotype nanobodies could mimic the antigen on amino acids and structures. Importantly, as more and more neutralization mAbs against different pathogens are prepared, anti-idiotype nanobody vaccines can be easily produced against the disease with our strategy, especially for dangerous pathogens.IMPORTANCEAnti-idiotype vaccines utilize idiotype-anti-idiotype network theory, eliminating the need for external antigens as vaccine candidates. Especially for dangerous pathogens, they were safer because they did not contact the live pathogenic microorganisms. However, developing anti-idiotype vaccines with traditional monoclonal and polyclonal antibodies is complex and has a high failure rate. We present a novel, universal, simple, low-cost strategy for producing anti-idiotype vaccines with nanobody technology. Using a neutralization antibody against PCV2-Cap, a nanobody (Ab2) was successfully produced and could mimic the neutralizing epitope of PCV2-Cap. The nanobody can induce protective immune responses against PCV2 infection in mice and pigs. It highlighted that the anti-idiotype vaccine using nanobody has a very good application in the future, especially for dangerous pathogens.


Asunto(s)
Infecciones por Circoviridae , Circovirus , Anticuerpos de Dominio Único , Vacunas Virales , Animales , Humanos , Ratones , Proteínas de la Cápside , Infecciones por Circoviridae/prevención & control , Infecciones por Circoviridae/veterinaria , Epítopos , Porcinos , Vacunas Virales/química , Vacunas Virales/inmunología
2.
J Clin Microbiol ; 59(12): e0158021, 2021 11 18.
Artículo en Inglés | MEDLINE | ID: mdl-34524888

RESUMEN

Porcine reproductive and respiratory syndrome virus (PRRSV) infection causes considerable economic loss to the global pig industry. Efficient detection assay is very important for the prevention of the virus infection. Nanobodies are the advantages of small molecular weight, simple genetic engineering, and low production cost for promising diagnostic application. In this study, to develop a nanobody-based competitive ELISA (cELISA) for specifically detecting antibodies against PRRSV, three nanobodies against PRRSV-N protein were screened by camel immunization, library construction, and phage display. Subsequently, a recombinant HEK293S cell line stably secreting nanobody-horseradish peroxidase (HRP) fusion protein against PRRSV-N protein was successfully constructed using the lentivirus transduction assay. Using the cell lines, the fusion protein was easily produced. Then, a novel cELISA was developed using the nanobody-HRP fusion protein for detecting antibodies against PRRSV in pig sera, exhibiting a cut-off value of 23.19% and good sensitivity, specificity, and reproducibility. Importantly, the cELISA specifically detect anti-genotype 2 PRRSV antibodies. The cELISA showed more sensitive than the commercial IDEXX ELISA kit by detecting the sequential sera from the challenged pigs. The compliance rate of cELISA with the commercial IDEXX ELISA kit was 96.4%. In addition, the commercial IDEXX ELISA kit can be combined with the developed cELISA for the differential detection of antibodies against genotype 1 and 2 PRRSV in pig sera. Collectively, the developed nanobody-based cELISA showed advantages of simple operation and low production cost and can be as an assay for epidemiological investigation of genotype 2 PRRSV infection in pigs and evaluation after vaccination.


Asunto(s)
Síndrome Respiratorio y de la Reproducción Porcina , Virus del Síndrome Respiratorio y Reproductivo Porcino , Animales , Anticuerpos Antivirales , Ensayo de Inmunoadsorción Enzimática , Genotipo , Síndrome Respiratorio y de la Reproducción Porcina/diagnóstico , Virus del Síndrome Respiratorio y Reproductivo Porcino/genética , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Porcinos
3.
Vet Res ; 52(1): 25, 2021 Feb 17.
Artículo en Inglés | MEDLINE | ID: mdl-33596995

RESUMEN

Porcine reproductive and respiratory syndrome virus (PRRSV) is a highly contagious virus that has led to enormous economic loss worldwide because of ineffective prevention and treatment. In view of their minimized size, high target specificity and affinity, nanobodies have been extensively investigated as diagnostic tools and treatments of many diseases. Previously, a PRRSV Nsp9-specific nanobody (Nb6) was identified as a PRRSV replication inhibitor. When it was fused with cell-penetrating peptide (CPP) TAT, Nb6-TAT could enter the cells for PRRSV suppression. However, delivery of molecules by CPP lack cell specificity and have a short duration of action. PRRSV has a tropism for monocyte/macrophage lineage, which expresses high levels of Fcγ receptors. Herein, we designed a nanobody containing porcine IgG Fc (Fcγ) to inhibit PRRSV replication in PRRSV permissive cells. Fcγ fused Nb6 chimeric antibody (Nb6-pFc) was assembled into a dimer with interchain disulfide bonds and expressed in a Pichia pastoris system. The results show that Nb6-pFc exhibits a well-binding ability to recombinant Nsp9 or PRRSV-encoded Nsp9 and that FcγR-mediated endocytosis of Nb6-pFc into porcine alveolar macrophages (PAM) was in a dose-dependent manner. Nb6-pFc can inhibit PRRSV infection efficiently not only by binding with Nsp9 but also by upregulating proinflammatory cytokine production in PAM. Together, this study proposes the design of a porcine IgG Fc-fused nanobody that can enter PRRSV susceptible PAM via FcγR-mediated endocytosis and inhibit PRRSV replication. This research reveals that nanobody-Fcγ chimeric antibodies might be effective for the control and prevention of monocyte/macrophage lineage susceptible pathogeneses.


Asunto(s)
Inmunoglobulina G/inmunología , Macrófagos Alveolares/virología , Virus del Síndrome Respiratorio y Reproductivo Porcino/inmunología , Receptores de IgG/fisiología , Anticuerpos de Dominio Único/inmunología , Proteínas no Estructurales Virales/inmunología , Animales , Síndrome Respiratorio y de la Reproducción Porcina/inmunología , Síndrome Respiratorio y de la Reproducción Porcina/prevención & control , Síndrome Respiratorio y de la Reproducción Porcina/virología , Virus del Síndrome Respiratorio y Reproductivo Porcino/fisiología , Anticuerpos de Dominio Único/química , Porcinos , Replicación Viral
4.
World J Surg ; 44(6): 1835-1843, 2020 06.
Artículo en Inglés | MEDLINE | ID: mdl-32052106

RESUMEN

BACKGROUND: Management errors during pre-hospital care, triage process and resuscitation have been widely reported as the major source of preventable and potentially preventable deaths in multiple trauma patients. Common tools for defining whether it is a preventable, potentially preventable or non-preventable death include the Advanced Trauma Life Support (ATLS®) clinical guideline, the Injury Severity Score (ISS) and the Trauma and Injury Severity Score (TRISS). Therefore, these surrogated scores were utilized in reviewing the study's trauma services. METHODS: Trauma data were prospectively collected and retrospectively reviewed from January 1, 2018, to December 31, 2018. All cases of trauma death were discussed and audited by the Hospital Trauma Committee on a regular basis. Standardized form was used to document the patient's management flow and details in every case during the meeting, and the final verdict (whether death was preventable or not) was agreed and signed by every member of the team. The reasons for the death of the patients were further classified into severe injuries, inappropriate/delayed examination, inappropriate/delayed treatment, wrong decision, insufficient supervision/guidance or lack of appropriate guidance. RESULTS: A total of 1913 trauma patients were admitted during the study period, 82 of whom were identified as major trauma (either ISS > 15 or trauma team was activated). Among the 82 patients with major trauma, eight were trauma-related deaths, one of which was considered a preventable death and the other 7 were considered unpreventable. The decision from the hospital's performance improvement and patient safety program indicates that for every trauma patient, basic life support principles must be followed in the course of primary investigations for bedside trauma series X-ray (chest and pelvis) and FAST scan in the resuscitation room by a person who meets the criteria for trauma team activation recommended by ATLS®. CONCLUSION: Mechanisms to rectify errors in the management of multiple trauma patients are essential for improving the quality of trauma care. Regular auditing in the trauma service is one of the most important parts of performance improvement and patient safety program, and it should be well established by every major trauma center in Mainland China. It can enhance the trauma management processes, decision-making skills and practical skills, thereby continuously improving quality and reducing mortality of this group of patients.


Asunto(s)
Traumatismo Múltiple/mortalidad , Mejoramiento de la Calidad , Adolescente , Adulto , Atención de Apoyo Vital Avanzado en Trauma , Anciano , Femenino , Humanos , Puntaje de Gravedad del Traumatismo , Masculino , Auditoría Médica , Persona de Mediana Edad , Traumatismo Múltiple/terapia , Seguridad del Paciente , Estudios Retrospectivos , Adulto Joven
5.
Zhongguo Zhong Yao Za Zhi ; 38(20): 3432-7, 2013 Oct.
Artículo en Zh | MEDLINE | ID: mdl-24490548

RESUMEN

In natural conditions, fully ripe Cynomorium songaricum seeds parasitize in Nitraria tangutorum or N. sphaerocarpa or N. sibirica or Zygophyllum xanthoxylom and Peganum harmala, were used in this study to research the morphological characteristics, embryo rate, seed viability, 1 000-grain weight, purity, water content and the seeds of different host parasitic relationship with each other. The results showed that the morphology, color and surface characteristics of the C. songaricum seeds are very similar in different hosts. According to the seed morphology can not be judged on its host. For the host to N. tangutorum or Peganum harmala or N. sibirica, we should choose the round hole screen less than 0.923 1 mm and larger than 1.066 2 mm to cleaning seeds. For the C. songaricum seeds parasitic in N. sphaerocarpa, the choice of slightly less than 0.926 1 mm and larger than 0.985 3 mm round hole screen to cleaning. For the parasitic seeds in Z. xanthoxylom, less than 0.751 3 mm and slightly larger than 1.035 3 mm round hole screen could be used. Highy significant correlation was found among the morphological indexes in C. songaricum seeds (P < 0.01). Morphological indexes and 1 000-grain weight were significantly correlated (0.01 < P < 0.05), but with the seed viability and the embryo rate were not found significant correlation. Grain weight is not related with the seed viability and the Fully mature C. songaricum seed viability is high and water content is low. The difference of the habitats and the host plants should be considered in the seed quality assessment and classification. The C. songaricum seeds on host plants are not selective, and the C. songaricum seeds from the host plants could be parasitized in other host plants.


Asunto(s)
Cynomorium/fisiología , Especificidad del Huésped , Magnoliopsida/fisiología , China , Cynomorium/química , Cynomorium/crecimiento & desarrollo , Ecosistema , Semillas/química , Semillas/crecimiento & desarrollo , Semillas/fisiología
6.
Talanta ; 259: 124564, 2023 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-37080074

RESUMEN

Drug resistance is a worldwide health care crisis which impedes disease treatment and increases financial burden, especially for its multifactorial nature and high complexity. Herein, we developed a multiparametric approach to visualize and detect drug resistance in living cancer cells, through the combination of DNA-templated covalent protein labeling strategy and fluorescent resonance energy transfer technique. Gefitinib resistance in non-small cell lung cancer caused by mesenchymal-epidermal transition factor (Met) overexpression and hyperactivation was investigated as a proof-of-concept. Unlike the traditional single-factor investigation, the proposed approach evaluated the contribution of three important parameters towards the resistance, including the changes of Met expression level, the homodimerization of Met with itself and the heterodimerization of Met with epidermal growth factor receptor (EGFR). A multiple regression model based on these three parameters was tentatively established for evaluation of the resistance level of laboratory-developed resistant cells and evaluation of the resistance level of patient-derived cells. Such an approach facilitates a quick identification of a drug resistance, to evaluate not only the resistance level but also the resistance mechanism.


Asunto(s)
Antineoplásicos , Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Quinazolinas/uso terapéutico , Transducción de Señal , Resistencia a Antineoplásicos , Línea Celular Tumoral , Inhibidores de Proteínas Quinasas/farmacología , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Mutación
7.
Ann Transl Med ; 7(22): 698, 2019 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-31930099

RESUMEN

Mesh infection after large incisional ventral hernia repair is a clinical dilemma in abdominal wall hernia surgery. It is believed foreign material should be removed but it causes secondary trauma to the abdominal wall tissue and might be associated with a higher risk of complications. Currently, there is no consensus on mesh-preservation treatment in cases of mesh infection after hernia repair in general. Herein we present the case of a 27-year-old male who recovered well from mesh infection after large incisional ventral hernia repair by mesh-preservation approach. The path to success is choice of material of prosthetic mesh; surgical approach of hernia repair, sufficient wound irrigation and drainage, and acquiring sterility of the mesh surface by wound care techniques such as local iodophor packing and vacuum sealing drainage. Clinical cohorts are needed to verify the feasibility of mesh-preservation treatment of mesh infection after large incisional hernia repair.

8.
Front Microbiol ; 10: 2313, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31649651

RESUMEN

Prevention and control of infection by porcine reproductive and respiratory syndrome virus (PRRSV) remains a challenge, due to our limited understanding of the PRRSV invasion mechanism. Our previous study has shown that PRRSV glycoprotein GP5 interacts with MYH9 C-terminal domain protein (PRA). Here we defined that the first ectodomain of GP5 (GP5-ecto-1) directly interacted with PRA and this interaction triggered PRA and endogenous MYH9 to form filament assembly. More importantly, MYH9 filament assembly was also formed in GP5-ecto-1-transfected MARC-145 cells. Notably, PRRSV infection of MARC-145 cells and porcine alveolar macrophages also induced endogenous MYH9 aggregation and polymerization that were required for subsequent PRRSV internalization. Moreover, overexpression of S100A4, a MYH9-specific disassembly inducer, in MARC-145 cells significantly resulted in diminished MYH9 aggregation and marked inhibition of subsequent virion internalization and infection by both PRRSV-1 and PRRSV-2 isolates. The collective results of this work reveal a novel molecular mechanism employed by MYH9 that helps PRRSV gain entry into permissive cells.

9.
Ann Transl Med ; 6(16): 324, 2018 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-30363998

RESUMEN

Laparoscopic low anterior resection (LAR) with sphincter preservation for ultra-low rectal cancer is always a challenging operation in colorectal surgery. To achieve negative margins, reducing the difficulty and risks of the procedure are major goals for us. The marker meeting approach we reported can help to accomplish this goal. The key technique for the marker meeting approach is to ensure a clear distal margin in a low resection of the rectum by transanal dissection. This procedure allows access to the space around the distal rectum and mesorectum and to pack the gauzes in the distal part of the space as a landmark. Routine laparoscopic LAR was performed to dissect the space until reaching the gauzes packed above and achieve complete mobilization of the rectum and mesorectum. This surgical procedure is simpler and reduces the difficulty of the operation. Therefore, it is expected to reduce the risk of surgery-related complications and positive margins and is suitable to be widely applied and extended in clinical practice. The short-term and long-term clinical outcomes of the marker meeting approach need more research in large samples.

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