ABSTRACT
Brucellosis is an infectious disease that brings enormous economic burdens for developing countries. The Brucella melitensis (B. melitensis) M5-90 vaccine strain (M5-90) has been used on a large scale in China, but may cause abortions if given to pregnant goats or sheep subcutaneously during the late stages of gestation. Moreover, the vaccine M5-90 cannot differentiate natural from vaccinated infection. Therefore, a safer and more potent M5-90 vaccine is required. In this study, a vjbR mutant of M5-90 (M5-90ΔvjbR) was constructed and overcame these drawbacks. M5-90ΔvjbR strain showed reduced survival capability in murine macrophages (RAW 264.7) and BALB/c mice and induced high protective immunity in mice. In addition, M5-90ΔvjbR induced an anti-Brucella-specific immunoglobulin G (IgG) response and stimulated the expression of gamma interferon (INF-γ) and interleukin-4 (IL-4) in vaccinated mice. Furthermore, M5-90ΔvjbR induced IgG response and stimulated the secretion of IFN-γ and IL-4 in immunized sheep. Moreover, the VjbR antigen allowed serological differentiation between infected and vaccinated animals. These results suggest that M5-90ΔvjbR is an ideal live attenuated and efficacious live vaccine candidate against B. melitensis 16â¯M infection.
Subject(s)
Bacterial Proteins/genetics , Bacterial Proteins/immunology , Brucella Vaccine/immunology , Brucella melitensis/immunology , Brucellosis/prevention & control , Disease Models, Animal , Animals , Antibodies, Bacterial/immunology , Bacterial Proteins/administration & dosage , Brucella Vaccine/administration & dosage , Brucella Vaccine/genetics , Brucella melitensis/genetics , Brucellosis/immunology , Brucellosis/microbiology , Drug Evaluation, Preclinical , Female , Gene Deletion , Humans , Interferon-gamma/genetics , Interferon-gamma/immunology , Interleukin-4/genetics , Interleukin-4/immunology , Mice , Mice, Inbred BALB C , Sequence Deletion , Vaccines, Attenuated/administration & dosage , Vaccines, Attenuated/genetics , Vaccines, Attenuated/immunologyABSTRACT
Aquaporin 3 (AQP3) has been shown to be low in the amnion and chorion tissues of patients with oligohydramnios and that S. miltiorrhiza, a Chinese herbal medicine, results in increased AQP3 in human amniotic epithelial cells (hAECs). Here, we provide evidence for the involvement of the JNK pathway in AQP3 regulation in isolated oligohydramnios tissues in vitro, in hAECs derived from normal amniotic fluid and fluid from patients with isolated oligohydramnios. Phosphorylation of JNK was suppressed by pretreatment of cells with JNK-specific inhibitor (SP600125) and was up-regulated by S. miltiorrhiza; S. miltiorrhiza combined with SP600125 prevented SP600125-induced down-regulation of phospho-JNK both in normal amniotic fluid volume and in isolated oligohydramnios. In isolated oligohydramnios, AQP3 expression was signiï¬cantly suppressed by SP600125 in a concentration- and time-dependent mannner, while its expression was up-regulated by S. miltiorrhiza. S. miltiorrhiza combined with SP600125 inhibited the increased expression of AQP3 relative to the S. miltiorrhiza treated group. Together, the data suggest that c-jun N-terminal kinase (JNK) pathway unerlies the regulation of AQP3 by S. miltiorrhiza amnion and chorion tissues.
Subject(s)
Aquaporin 3/metabolism , MAP Kinase Signaling System , Oligohydramnios/metabolism , Adult , Amnion/drug effects , Amnion/metabolism , Amniotic Fluid/drug effects , Anthracenes/administration & dosage , Case-Control Studies , Cells, Cultured , Drugs, Chinese Herbal/administration & dosage , Epithelial Cells/drug effects , Epithelial Cells/metabolism , Female , Humans , MAP Kinase Signaling System/drug effects , Oligohydramnios/drug therapy , Pregnancy , Salvia miltiorrhiza , Young AdultABSTRACT
OBJECTIVE: To explore the role of Compound Danshen Injection (CDI) in regulating the expression of aquaporin 3 (AQP3) in human amnion epithelium cells (hAECs), and to study the relation between c-Jun N-terminal kinase (JNK) signal pathway and AQP3. METHODS: hAECs were isolated and primarily cultured from term pregnancy with normal amniotic fluid volume and from term pregnancy with oligohydramnios, and then hAECs were further divided into four groups, i.e., the blank control group (A), the SP600125 group (B), the CDI group (C), and the SP600125 +CDI group (D). The cell viability was measured by cell counting kit-8 assay (CCK-8). The expression of total JNK, phosphorylated JNK, and AQP3 were determined by Western blot. RESULTS: (1) In hAECs with normal AFV or with oligohydramnios: There was no statistical difference in the cell viability or the expression of total JNK among the 4 groups (P > 0.05). But there was statistical difference in the expression of p-JNK (P < 0.05). Compared with A group, the expression of p-JNK was obviously down-regulated in B group, but obviously up-regulated in C group (P < 0.05). The expression of p-JNK was significantly lower in D group than in C group, but higher than that in A group or B group (P < 0.05).The AQP3 expression in the hAECs with normal amniotic fluid volume of C group and D group were higher than that in the A group (P < 0.05). However, there was no statistical difference in the AQP3 expression between C group and D group (P > 0.05). In hAECs with oligohydramnios, the expression of AQP3 obviously decreased in B group, but up-regulated in C group (both P < 0.05). The expression of AQP3 was lower in D group than in C group, but higher than in B group (P < 0.05). CONCLUSION: CDI could regulate the AQP3 expression in hAECs with oligohydramnios via activating the JNK signal pathway.