Purified Immunoglobulin F(ab′) 2 Protects Mice and Rhesus Monkeys against Lethal Ricin Intoxication

Ricin is a highly toxic ribosome-inactivating lectin derived from castor beans. To date, no antidote is available to treat ricin-poisoned patients, and the development of a safe and effective antidote is urgently needed. First, ricin was prepared and used to construct a mouse model and a rhesus monkey model of ricin intoxication. Second, pepsin-digested F(ab′) 2 fragments of serum IgG from horses injected with Freund’s-adjuvanted purified ricin were prepared. Third, the protective efficacy was evaluated in mouse and rhesus monkey models of lethal ricin intoxication. The purity quotient of the prepared ricin and F(ab′) 2 fragments exceeded 90% and 85% in the mouse and monkey models, respectively. The LD 50 of ricin in mice and rhesus monkeys was 2.7 and 9 μg/kg, respectively. A quantity of 6.25 and 1.85 mg/kg F(ab′) 2 was sufficient to treat lethal ricin intoxication in the mice and rhesus monkeys, respectively. Finally, the effect of this therapeutic antibody on peripheral blood immune cells was examined by analysis of peripheral blood immune cells through single cell sequencing. The underlying mechanism was found to involve restraining neutrophil activation, proliferation, and differentiation. Purified F(ab′) 2 fragments administered with needle-free devices fully protect mice and rhesus monkeys against lethal doses of ricin intoxication

Development and Efficacy Evaluation of an SP01-adjuvanted Inactivated Escherichia Coli Mutant Vaccine Against Bovine Coliform Mastitis

Escherichia coli ( E. coli ) is one of the most common pathogens causing clinical mastitis in cattle, but no vaccine is available to prevent this disease in China. Therefore, development of an E. coli vaccine against bovine clinical mastitis is urgently needed. The candidate vaccine (Ch-O111-1) and challenge (LZ06) strains were screened from milk samples of cows with clinical mastitis. To extend the cross-protection of the Ch-O111-1 strain, we deleted the galE gene fragment of the Ch-O111-1 strain through homologous recombination between the Ch-O111-1 strain and pCVD442/ΔgalE plasmid, which was identified through conventional methods, including PCR, SDS-PAGE and sequencing. The Ch-O111-1/ΔgalE (Z9) strain was characterized by extensive cross-reactivity and attenuated virulence. We prepared inactivated Z9 vaccines with different adjuvants. Immunization of inactivated Z9 antigen induced adjuvant-, dosage- and inoculation time-dependent antibody titers in cows and mice. Furthermore, immunization with SP01-adjuvanted inactivated Z9 vaccine protected cows against severe clinical mastitis caused by LZ06 and protected mice against death due to LZ06. An SP01-adjuvanted inactivated Z9 vaccine was successfully developed and found to protect cows against severe mastitis caused by Escherichia coli

Construction and Evaluation of the Brucella Double Gene Knock-out Vaccine Strain MB6 Δbp26ΔwboA (RM6)

Brucellosis is a serious zoonotic infection worldwide. To date, vaccination is the most effective measure against brucellosis. This study was aimed at obtaining a vaccine strain that has high protective efficacy and low toxicity, and allows vaccination to be differentiated from infection. Using homologous recombination, we constructed a double gene-deletion Brucella strain MB6 Δbp26ΔwboA (RM6) and evaluated its characteristics, safety and efficacy. The RM6 strain had good proliferative ability and stable biological characteristics in vivo and in vitro. Moreover, it had a favorable safety profile and elicited specific immune responses in mice and sheep. The RM6 strain may have substantial practical application value

Preparation of Equine Immunoglobulin F(ab′) 2 against Smallpox and Evaluation of its Immunoprotective Effect

Smallpox, a severe infectious disease caused by the smallpox virus, causes a death rate as high as 30% within 15-20 days after infection. Therefore, development of anti-Smallpox product as a strategic reserve is urgently needed. We prepared and tested pepsin-digested F(ab′) 2 fragments of serum IgG from horses. Transmission electron microscopy indicated that the purified virus showed morphology consistent with VVTT. The titer was above 1.0 × 10 7 PFU/mL. The purity of the antigen exceeded 90%, according to HPLC. After purification and cleavage, the yield of the purified product F(ab′) 2 was approximately 1.3%, its purity exceeded 90%, and the neutralizing antibody titer exceeded 1:3200. F(ab′) 2 fragments had good preventive and therapeutic effects in mice at antibody doses of 5.2 mg/mL and 2.6 mg/mL. The viral loads of the drug-treated mice were suppressed to varying degrees, and the higher dose groups (5.2 and 2.6 mg/mL) showed a 2-3 fold lower viral load than that in the control group. A process for producing equine immunoglobulin F(ab′) 2 against VVTT was established. The prepared horse anti-smallpox immunoglobulin product had good neutralizing antibody effects on VVTT. The highly purified preparation may serve as a potential candidate for smallpox treatment

H₂O₂/Glucose Sensor Based on a Pyrroloquinoline Skeleton-Containing Molecule Modified Gold Cavity Array Electrode

H2O2-related metabolites are essential indicators in clinical diagnosis because the accumulation of such reactive oxygen species could cause the risk of cardiovascular disease. Herein, we reported an electrochemical sensor to determine H2O2 and glucose. The pyrroloquinoline skeleton containing molecules (PQT) were used as the electrocatalyst and the gold cavity array (GCA) electrodes as the supporting electrode. The GCA electrode was fabricated by electrodeposition using high-ordered two-dimensional polystyrene spheres as the template. The strong absorbability of iodide ions (I−) displaced adventitious materials from the metal surface and the I− monolayer was subsequently removed by electrochemical oxidation to get a clean electrode surface. PQT molecules were firmly immobilized on the GCA electrode and performed an excellent electrocatalytic effect on H2O2/glucose detection, manifested by a small overpotential and a significantly increased reduction current. A good linear correlation was observed over a wide range of 0.2 μmol/L–1.0 mmol/L with the limit of detection of 0.05 μmol/L. Moreover, the sensor can realize sensitive, accurate, and the highly selective detection of actual samples, proving its application prospect in clinical diagnosis

miR-10a rejuvenates aged human mesenchymal stem cells and improves heart function after myocardial infarction through KLF4

Abstract Background Aging is one of the key factors that regulate the function of human bone marrow mesenchymal stem cells (hBM-MSCs) and related changes in microRNA (miRNA) expression. However, data reported on aging-related miRNA changes in hBM-MSCs are limited. Methods We demonstrated previously that miR-10a is significantly decreased in aged hBM-MSCs and restoration of the miR-10a level attenuated cell senescence and increased the differentiation capacity of aged hBM-MSCs by repressing Krüpple-like factor 4 (KLF4). In the present study, miR-10a was overexpressed or KLF4 was downregulated in old hBM-MSCs by lentiviral transduction. The hypoxia-induced apoptosis, cell survival, and cell paracrine function of aged hBM-MSCs were investigated in vitro. In vivo, miR-10a-overexpressed or KLF4-downregulated old hBM-MSCs were implanted into infarcted mouse hearts after myocardial infarction (MI). The mouse cardiac function of cardiac angiogenesis was measured and cell survival of aged hBM-MSCs was investigated. Results Through lentivirus-mediated upregulation of miR-10a and downregulation of KLF4 in aged hBM-MSCs in vitro, we revealed that miR-10a decreased hypoxia-induced cell apoptosis and increased cell survival of aged hBM-MSCs by repressing the KLF4–BAX/BCL2 pathway. In vivo, transplantation of miR-10a-overexpressed aged hBM-MSCs promoted implanted stem cell survival and improved cardiac function after MI. Mechanistic studies revealed that overexpression of miR-10a in aged hBM-MSCs activated Akt and stimulated the expression of angiogenic factors, thus increasing angiogenesis in ischemic mouse hearts. Conclusions miR-10a rejuvenated aged hBM-MSCs which improved angiogenesis and cardiac function in injured mouse hearts

NLRP6 Induces Lung Injury and Inflammation Early in Brucella and Influenza Coinfection

(1) Background: With the resurgence of brucellosis epidemics in China in recent years, the chances of a brucella coinfection with other common respiratory pathogens, such as the influenza virus, have increased dramatically. However, little is known about the pathogenicity or the mechanisms of brucella and influenza coinfections. (2) Methods: To clarify the interventions in the early stages of lung damage due to brucella and influenza coinfections, we evaluated the effect of the coinfection on disease progression and mortality using a coinfection model in WT mice and NLRP6−/− mice, and we verified the function of NLRP6 in infection and proinflammation. (3) Results: The coinfection induced significant respiratory symptoms, weight loss, and a high mortality rate in WT mice. Influenza in the coinfection group significantly increased brucella proliferation in a synergistic manner. Meanwhile, a histological examination showed severe lung tissue destruction and excessive inflammatory responses in coinfected WT animals, and the expression of NLRP6 and IL-18 was dramatically increased in the lung tissues. Furthermore, NLRP6 deletion attenuated lung injuries and inflammation, a reduced bacterial load, and decreased IL-18 protein expression. (4) Conclusions: Our findings indicated that NLRP6 plays a critical role and might be a promising potential therapeutic target for brucella–influenza coinfections

Additional file 7: of miR-10a rejuvenates aged human mesenchymal stem cells and improves heart function after myocardial infarction through KLF4

Figure S6. Downregulation of miR-10a or overexpression of KLF4 in old hBM-MSCs increased apoptotic gene expression. Quantification of mRNA expression of BAX and PUMA (pr-apoptotic), BCL2 and MCL1 (antiapoptotic) in O, O-anti10a and O-KLF4 hBM-MSCs after culture for 72 h under hypoxia conditions. n = 6/group. Mean ± SD. *P < 0.05 (PDF 38 kb

Additional file 1: of miR-10a rejuvenates aged human mesenchymal stem cells and improves heart function after myocardial infarction through KLF4

Table S1. qRT-PCR primer and miRNA RT primer sequences (DOCX 19 kb

Additional file 5: of miR-10a rejuvenates aged human mesenchymal stem cells and improves heart function after myocardial infarction through KLF4

Figure S4. Overexpression of miR-10a in both young and old hBM-MSCs decreased hypoxia-induced apoptosis and increased cell survival. miR-10a transduced into young (Y-10a) and old (O-10a) hBM-MSCs by lentiviral vector. Control vector-transduced young hBM-MSCs (Y-c) and old hBM-MSCs (O-c) served as controls. Cells cultured for 72 h under hypoxia conditions. (A) Cell apoptosis assayed by TUNEL staining. Percentage of apoptotic cells (TUNEL+) quantified in Y-c, Y-10a, O-c, and O-10ahBM-MSCs. (B) Cell survival evaluated in Y-c, Y-10a, O-c, and O-10a hBM-MSCs. n = 6/group. Mean ± SD. *P < 0.05, Y-c vs Y-10a; #P < 0.05, O-c vs O-10a (PDF 94 kb