RESUMO
OBJECTIVE: To observe the effects of neural stem cells (NSC) plus self-assembly isoleucine-lysine-valine-alanine-valine (IKVAV) nanofiber gel transplantation on the promotion of function recovery of spinal cord injury (SCI) in rats. METHODS: A total of 230 SD rats were randomized into gel, NSC, NSC plus self-assembly IKVAV nanofiber gel transplantation, normal saline and sham-operation groups. Function repair was evaluated by bundle branch block (BBB) score, immunofluorescence and Western blot respectively at Day 1, 3, 5, 7, 14, 28, 56 and 92 post-operation. RESULTS: There were statistically significant differences among bundle branch block (BBB) scores of different treatment groups (P < 0.01). Moreover, statistical significance existed between each treatment group and combined transplantation group (P = 0.000). The expression of glial fibrillary acidic protein in combined transplantation group (rats with spinal injury) was lower than that in other treatment groups (except for sham operation) and the expression of NF-200 in this group was higher than that in other treatment groups (except for sham operation). Significant differences existed in the expressions of brain-derived neurotrophic factor and nerve growth factor between combined transplantation and other treatment groups (P < 0.01). CONCLUSION: Transplantation with IKVAV nanofiber gel, NSC and NSC plus self-assembly IKVAV nanofiber gel may promote the repair of SCI in rats. But the method of NSC plus self-assembly IKVAV nanofiber gel is more effective.
Assuntos
Laminina/uso terapêutico , Células-Tronco Neurais/transplante , Fragmentos de Peptídeos/uso terapêutico , Traumatismos da Medula Espinal/cirurgia , Animais , Géis/uso terapêutico , Masculino , Nanofibras , Ratos , Ratos Sprague-Dawley , Recuperação de Função FisiológicaRESUMO
Echinococcus multilocularis larvae, predominantly located in the liver, cause a tumor-like parasitic disease, alveolar echinococcosis (AE), that is characterized by increased infiltration of various immune cells, including macrophages, around the lesion that produces an "immunosuppressive" microenvironment, favoring its persistent infection. However, the role of hepatic macrophages in the host defense against E. multilocularis infection remains poorly defined. Using human liver tissues from patients with AE and a hepatic experimental mouse model of E. multilocularis, we investigated the phenotype and function of hepatic macrophages during the parasite infection. In the present study, we found that a large number of CD68+ macrophages accumulated around the metacestode lesion in the liver of human AE samples and that both S100A9+ proinflammatory (M1 phenotype) and CD163+ anti-inflammatory (M2 phenotype) macrophages were significantly higher in close liver tissue (CLT) than in distant liver tissue (DLT), whereas M2 macrophages represent the dominant macrophage population. Furthermore, E. multilocularis-infected mice exhibited a massive increase in macrophage (F4/80+) infiltration in the liver as early as day 5, and the infiltrated macrophages were mainly monocyte-derived macrophages (CD11bhi F4/80int MoMFs) that preferentially differentiated into the M1 phenotype (iNOS+) at the early stage of E. multilocularis infection and then polarized to anti-inflammatory macrophages of the M2 phenotype (CD206+) at the chronic stage of infection. We further showed that elimination of macrophages by treatment of mice with clodronate-liposomes before E. multilocularis infection impaired worm expulsion and was accompanied by a reduction in liver fibrosis, yielding a high parasite burden. These results suggest that hepatic macrophages may play a dual role in the establishment and development of E. multilocularis metacestodes in which early larvae clearance is promoted by M1 macrophages while persistent metacestode infection is favored by M2 macrophages.
Assuntos
Equinococose , Echinococcus multilocularis/imunologia , Estágios do Ciclo de Vida/imunologia , Fígado , Macrófagos , Animais , Equinococose/imunologia , Equinococose/parasitologia , Equinococose/patologia , Feminino , Humanos , Fígado/imunologia , Fígado/parasitologia , Fígado/patologia , Macrófagos/imunologia , Macrófagos/parasitologia , Macrófagos/patologia , CamundongosRESUMO
OBJECTIVE: To isolate mesenchymal stem cells (MSCs) from second-trimester amniotic fluid using an improved two-stage culture protocol, and to induce these MSCs into neuron-like cells. METHODS: An improved two-stage culture protocol for MSCs from amniotic fluid was developed. MSCs from amniotic fluid were induced to differentiate with beta-mercaptoethanol into neuron-like cells. Flow cytometry, reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry were employed for analysis of the phenotypic characteristics of the cultured MSCs from amniotic fluid. RESULTS: MSCs from amniotic fluid were successfully isolated, cultured and enriched without interfering with the routine process of fetal karyotyping. Flow cytometry analysis showed that they were positive for CD44, CD29, and CD105, but negative for CD45, CD34, or human leucocyte antigen-DR (HLA-DR). Importantly, a subpopulation of transcription factor Oct-4 positive cells could be detectable in the cultured MSCs from amniotic fluid. Moreover, MSCs from amniotic fluid obtained by two-stage culture protocol showed higher proliferation rate [(15.0+/-2.3)% vs. (10.0+/-1.8)%] and higher Oct-4 positive cell rate [(1.2+/-0.3)% vs. (0.9+/-0.2)%, both P<0.05]. After the induction, MSCs from amniotic fluid displayed processes that formed extensive networks. Positive cells for neurone specific enolase (NSE) constituted (54.76+/-3.65)%, and glial fibrillary acidic protein (GFAP) (36.28+/-4.27)%, respectively. CONCLUSION: We demonstrate that human pluripotent MSCs are present in second-trimester amniotic fluid. The two-stage culture protocol could be a kind of simple one with high performance and it does not interfere with the routine fetal karyotyping. MSCs from amniotic fluid can be induced to differentiate into neuron-like cells in vitro by beta-mercaptoethanol in optimal medium.
Assuntos
Líquido Amniótico/citologia , Técnicas de Cultura de Células/métodos , Células-Tronco Mesenquimais/citologia , Diferenciação Celular , Células Cultivadas , Células-Tronco Embrionárias/citologia , Humanos , Neurônios/citologiaRESUMO
BACKGROUND: Larvae of Echinococcus granulosus (sensu lato) dwell in host organs for a long time but elicit only a mild inflammatory response, which indicates that the resolution of host inflammation is necessary for parasite survival. The recruitment of alternatively activated macrophages (AAMs) has been observed in a variety of helminth infections, and emerging evidence indicates that AAMs are critical for the resolution of inflammation. However, whether AAMs can be induced by E. granulosus (s.l.) infection or thioredoxin peroxidase (TPx), one of the important molecules secreted by the parasite, remains unclear. METHODS: The activation status of peritoneal macrophages (PMs) derived from mice infected with E. granulosus (sensu stricto) was analyzed by evaluating the expression of phenotypic markers. PMs were then treated in vivo and in vitro with recombinant EgTPx (rEgTPx) and its variant (rvEgTPx) in combination with parasite excretory-secretory (ES) products, and the resulting activation of the PMs was evaluated by flow cytometry and real-time PCR. The phosphorylation levels of various molecules in the PI3K/AKT/mTOR pathway after parasite infection and antigen stimulation were also detected. RESULTS: The expression of AAM-related genes in PMs was preferentially induced after E. granulosus (s.s.) infection, and phenotypic differences in cell morphology were detected between PMs isolated from E. granulosus (s.s.)-infected mice and control mice. The administration of parasite ES products or rEgTPx induced the recruitment of AAMs to the peritoneum and a notable skewing of the ratio of PM subsets, and these effects are consistent with those obtained after E. granulosus (s.s.) infection. ES products or rEgTPx also induced PMs toward an AAM phenotype in vitro. Interestingly, this immunomodulatory property of rEgTPx was dependent on its antioxidant activity. In addition, the PI3K/AKT/mTOR pathway was activated after parasite infection and antigen stimulation, and the activation of this pathway was suppressed by pre-treatment with an AKT/mTOR inhibitor. CONCLUSIONS: This study demonstrates that E. granulosus (s.s.) infection and ES products, including EgTPx, can induce PM recruitment and alternative activation, at least in part, via the PI3K/AKT/mTOR pathway. These results suggest that EgTPx-induced AAMs might play a key role in the resolution of inflammation and thereby favour the establishment of hydatid cysts in the host.
Assuntos
Echinococcus granulosus/imunologia , Macrófagos Peritoneais/imunologia , Proteína Oncogênica v-akt/metabolismo , Peroxirredoxinas/imunologia , Fosfatidilinositol 3-Quinases/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Equinococose/parasitologia , Echinococcus granulosus/enzimologia , Feminino , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Peroxirredoxinas/farmacologia , Fenótipo , Fosforilação , Transdução de Sinais , Organismos Livres de Patógenos EspecíficosRESUMO
Alveolar echinococcosis (AE) is a prevalent epidemic in the northern hemisphere, especially in central Europe and western China. Serum diagnosis is important for patients with AE, especially during the first screening. The present study purified the recombinant Em18-GST (rEm18-GST), and detected its diagnostic performance in human alveolar echinococcosis patients of Xinjiang, China with immunoblotting (IB) and enzyme-linked immunosorbent assay (ELISA). Serum samples were collected from 50 patients with AE, 222 patients with cystic echinococcosis (CE), 158 patients with other unrelated infections and 106 healthy individuals. The IB results showed that serum samples of 47 patients with AE and 12 patients with CE were rEm18-positive. However, only one sample from patients with cancer showed a cross-reaction with rEm18 in IB. The overall sensitivity was 94%, and the total specificity was 96.58%. For the rEm18 results using ELISA, the sera of 46 patients with AE were positive, and the overall sensitivity was 92%. In conclusion, compared with imaging tools, including computed tomography, magnetic resonance imaging and positron emission tomography, rEm18 has considerable advantages for AE serodiagnosis.