The neurotoxicity of lead has been well established, and oxidative stress is strongly associated with lead‐induced neurotoxicity. Nrf2 is important for protection against oxidative stress in many disease models. We applied t‐BHQ, which is an Nrf2 activator, to investigate the possible role of Nrf2 in the protection against lead neurotoxicity. t‐BHQ significantly attenuated the oxidative stress in developmental rats by decreasing MDA level, as well as by increasing SOD activity and GSH content, in the hippocampus and frontal cortex. Furthermore, neuronal apoptosis was detected by Nissl staining, and Bax expression was inhibited in the t‐BHQ‐treated group. Results showed that t‐BHQ suppressed ROS production and caspase 3/7 activity but increased intracellular GSH content, in SH‐SY5Y cells under lead exposure. Moreover, in vivo and in vitro , t‐BHQ enhanced the nuclear translocation of Nrf2 and binding to ARE areas but did not induce Nrf2 transcription. These phenomena were confirmed using RT‐PCR, EMSA, Western blot, and immunofluorescence analyses. Subsequent upregulation of the expression of HO‐1, NQO1, and GCLC was observed. However, knockdown of Nrf2 or HO‐1 adversely affected the protective effects of t‐BHQ against lead toxicity in SH‐SY5Y cells. Thus, t‐BHQ can protect against lead neurotoxicity, depending on the Nrf2/HO‐1 pathway.
Abstract Background The use of stem cell-derived exosomes (Exos) as therapeutic vehicles is receiving increasing attention. Exosome administration has several advantages over cell transplantation, thus making exosomes promising candidates for large-scale clinical implementation and commercialization. However, exosome extraction and purification efficiencies are relatively low, and therapeutic heterogeneity is high due to differences in culture conditions and cell viability. Therefore, in this study, we investigated a priming procedure to enhance the production and therapeutic effects of exosomes from human umbilical cord mesenchymal stem cells (hucMSCs). After preconditioning hucMSCs with agonists/inhibitors that target the Wnt/β-catenin pathway, we assessed both the production of exosomes and the therapeutic efficacy of the optimized exosomes in the context of diabetic wound healing, hoping to provide a safer, more stable and more effective option for clinical application. Results The Wnt signalling pathway agonist CHIR99021 increased exosome production by 1.5-fold without causing obvious changes in the characteristics of the hucMSCs or the size of the exosome particles. Further studies showed that CHIR99021 promoted the production of exosomes by facilitating exocytosis. This process was partly mediated by SNAP25. To further explore whether CHIR99021 changed the cargo that was loaded into the exosomes and its therapeutic effects, we performed proteomic and transcriptomic analyses of exosomes from primed and control hucMSCs. The results showed that CHIR99021 significantly upregulated the expression of proteins that are associated with cell migration and wound healing. Animal experiments confirmed that, compared to control hucMSC-derived exosomes, CHIR99021-pretreated hucMSC-derived exosomes (CHIR-Exos) significantly accelerated wound healing in diabetic mice, enhanced local collagen deposition, promoted angiogenesis, and reduced chronic inflammation. Subsequent in vitro experiments confirmed that the CHIR-Exos promoted wound healing by facilitating cell migration, inhibiting oxidative stress-induced apoptosis, and preventing cell cycle arrest. Conclusions The Wnt agonist CHIR99021 significantly increased exosome secretion by hucMSCs, which was partly mediated by SNAP25. Notably, CHIR99021 treatment also significantly increased the exosomal levels of proteins that are associated with wound healing and cell migration, resulting in enhanced acceleration of wound healing. All of these results suggested that pretreatment of hucMSCs with CHIR99021 not only promoted exosome production but also improved the exosome therapeutic efficacy, thus providing a promising option for large-scale clinical implementation and commercialization. Graphical Abstract
To study the relaxation effects of tetrandrine on the corpus cavernosum tissue of rabbit in vitro.1. The fluctuation of the dose-response relaxation curves for the contraction of KCl induced by tetrandrine was observed with isolated rabbit corpus cavernosum tissue. 2. Isolated strips of rabbit corpus cavernosum tissue were precontracted with 10 mumol/L phenylephrine(PE). Relaxation in response to cumulative doses of tetrandrine was determined in the absence and presence of nitric oxide synthase inhibitor (L-NNA) and soluble guanylate cyclase inhibitor (methylthioninium).1. The dose-response curves of KCl were shifted to the right nonparallelly, and the maximal responses were depressed to (73.0 +/- 3.8)% and (41.5 +/- 3.4)%, respectively, in the presence of tetrandrine(10 mumol/L, 30 mumol/L). 2. On rabbit cavernosal muscle stripes precontracted with PE(10 mumol/L), increasing concentrations of tetrandrine (1 mumol/L, 10 mumol/L, 30 mumol/L and 100 mumol/L) showed dose dependent relaxation [(6.0 +/- 1.4)%, (21.3 +/- 2.2)%, (47.4 +/- 3.3)%, and (68.1 +/- 3.6)%, P < 0.01]. However, in the meantime, it was found that these relaxation effects were not affected by the presence of L-NNA and methylthioninium (P > 0.05).Tetrandrine was effective in relaxing rabbit corpus cavernosum tissue in vitro in a dose-dependent style. The mechanism might be related with its blocking effect on calcium channel, but not the NO-cGMP passage.
Currently, general immunosuppressive drugs are used to maintain tolerance to allografts. However, these drugs have a major drawback of rendering the patient susceptible to infections and other side effects like malignancy and drug related toxicities with an overall rejection of the organ at some point. Previous studies have shown that MHC-Ig dimers may suppress alloresponsive T cells in a donor specific manner in vitro. This work aimed to answer the question as to whether these dimers will surmount rejection through the direct mechanism of allorecognition by suppressing alloreactive CD8+ T cells. To do this, we first identified two mice models with a single mismatch at the MHC loci. We found and procured white albino NOD mice which happened to be transgenic for HLA-A2 and HLA-A24 molecules. We then constructed a human-mouse hybrid HLA-A2-Ig dimer by overlap-PCR to join parts of two different already cloned plasmids to form the full length HLA-A2β2α1α2murineα3 insert which was then cloned to pcDNA3.1 to form pcDNA3.1HLA-A2β2α1α2murineα3. The IgG2bFc region was added by restriction digestion and ligation to form the plasmid pcDNA3.1HLA-A2β2α1α2murineα3IgG2bFc. Sequencing was done and confirmed that the construction and cloning were successful. The plasmid pcDNA3.1HLA-A2β2α1α2murineα3IgG2bFc was then transfected by electroporation to J558L cells. Screening was done using G418 for 4 weeks in cell culture. We purified the dimer by affinity chromatography and then used ELISA to confirm expression of the dimer. The purified dimer was then used in 1-way MLC experiments where responder cells were mice cells expressing HLA-A24 molecules while stimulator cells were mice cells expressing HLA-A2 molecules. Cell samples were gated on anti-mouse CD3-PE/CY7, anti-mouse CD4-PE, and anti-mouse CD8-APC/CY7. Cell proliferation was analysed using CFSE. Our results showed that the proliferation of CD4+ T cells was inhibited in the presence of the dimer. This work is crucial for subsequent studies aiming to search for induction of donor specific tolerance.
Objective: To investigate the therapy effect of valsartan on oxidative stress and the formation of atherosclerosis of rabbit. Methods: An atherosclerotic rabbit model was established by feeding high cholesterol diet supplemented by bovine serum albumin injection bolus. The rabbits were randomly divided into the control, model, and valsartan treated group, six rabbits in each group. Blood samples were collected at the end of 8 weeks for examination of serum lipid levels and MDA levels; the aortas were harvested for histological morphometry analysis, vascular cell adhesion molecule-1 (VCAM-1) immunohistochemical analysis and in situ superoxide detection to reflect the activity of NAD(P)H oxidase. Results: Rabbits fed with high cholesterol diet showed higher serum lipids levels than those fed with normal diet(P0.01). Treatment with valsartan (10 mg/kg per day) did not alter serum lipids levels. But the serum MDA level and ratio of lesion to intima area reduced significantly compared with model group(P0.05). The expression of VCAM-1 decreased significantly in the valsartan treated group than in the model group (P0.05).In addition, in situ superoxide detection also show the markedly reduction of superoxide as a result of valsartan treatment. Conclusion: These results indicate that the valsartan treatment can reduce the atherosclerotic progression, the mechanisms of which may include the inhibiting the NAD(P)H oxidase activity to produce superoxide and the downregulating the expression of redox sensitive genes in the downstream, such as VCAM-1.
CD8(+) suppressor T cells have been demonstrated to provide protection of allografts from rejection. We previously reported that soluble peptide/HLA-A2 dimer shows peptide-specific inhibitory effects on alloresponse in a coculture of peptide-pulsed T2 cells with HLA-A2 negative lymphocytes in vitro. Here we found a subset of CD8(low)CD28(-) T cells that was induced in the dimer-treated coculture. Importantly, this population showed hyporesponsiveness to the alloantigen restimulation as well as alloantigen-specific suppression on alloreactive T cells in a cell-cell contact-dependent fashion. The suppressive mechanisms of CD8(low)CD28(-) T cells involved an elevated expression of membrane-bound TGF-β1, but not Foxp3, CTLA-4, or IL-10. Furthermore, an overrepresention of CD8(low)CD28(-) T cells was observed in the patients after allogeneic platelet transfusion and positively correlated with the elevated concentrations of plasma HLA class I antigens. Our findings demonstrated that soluble HLA-A2 dimer could efficiently induce the tolerant CD8(low)CD28(-) T cells with alloantigen-specific suppression on alloreactive T cells. This study might provide a new strategy for preparation of donor-specific suppressor T cells and represent an attractive alternative for induction of allograft tolerance.
Objective To investigate the effect of breviscapine on lung injury in children undergoing open heart surgery with cardiopulmonary bypass(CPB).Methods Forty-five ASA Ⅱ or Ⅲ children aged 3-65 months weighing 5-21 kg undergoing open heart surgery with CPB were randomly assigned to 3 groups ( n =15 each):control group (group C),low dose breviscapine group (group B1 ) and high dose breviscapine group (group B2).Normal saline 15 ml(group C),breviscapine 0.5 mg/kg (group B1 )or 1.0 mg/kg(group B2 ) were injected iv over 30min after anesthesia induction.Blood samples were taken before operation ( T0 ),at 30 min and 1 h of aortic unclamping (T1,T2 ),at 3 h and 6 h after operation (T3,T4 ) for determination of plasma procalcitonin (PCT)and neutrophil elastase(NE) concentrations.PaO2 and PaCO2 were recorded at T0,T3,T4 for caculation of oxygenation index (OI) and alveolo-arterial oxygen partial pressure difference (PA-a O2 ).Results There were no significant differences in OI and PA-a O2 among the 3 groups( P ＞ 0.05).Plasma concentration of PCT was higher at T1～4in 3 groups,and plasma concentration of NE was higher at T1 in group C than that at T0 ( P ＜ 0.01 ).Plasma concentrations of PCT and NE were lower in groups B1 and B2 than in group C ( P ＜ 0.01).There were no significant differences in plasma concentrations of PCT and NE between groups B1 and B2 ( P ＞ 0.05).Conclusion Breviscapine(0.5,1 mg/kg) can inhibite systemic inflammatory response and attenuate lung injury in children undergoing open heart surgery with CPB.
Key words:
Flavonoids; Cardiopulmonary bypass; Cardiac surgical procedures; Respiratory distress syndrome, adult; Child
-Di(2-ethylhexyl) phthalate (DEHP), a typical endocrine-disrupting chemical (EDC), is widely used as plasticizer. DEHP exposure in humans is virtually ubiquitous, and those undergoing certain medical procedures can be especially high. In this study, we investigated whether developmental DEHP exposure disrupted glucose homeostasis in the rat and whether this was associated with the early impairment in endocrine pancreas. Pregnant Wistar rats were administered DEHP (1.25 and 6.25 mg·kg(-1)·day(-1)) or corn oil throughout gestation and lactation by oral gavage. Body weight, glucose and insulin tolerance, and β-cell morphometry and function were examined in offspring during the growth. In this study, developmental DEHP exposure led to abnormal β-cell ultrastructure, reduced β-cell mass, and pancreatic insulin content as well as alterations in the expression of genes involved in pancreas development and β-cell function in offspring at weaning. At adulthood, female DEHP-exposed offspring exhibited elevated blood glucose, reduced serum insulin, impaired glucose tolerance, and insulin secretion. Male DEHP-exposed offspring had increased serum insulin, although there were no significant differences in blood glucose at fasting and during glucose tolerance test. In addition, both male and female DEHP-exposed offspring had significantly lower birth weight and maintained relatively lower body weight up to 27 wk of age. These results suggest that developmental exposure to DEHP gives rise to β-cell dysfunction and the whole body glucometabolic abnormalities in the rat. DEHP exposure in critical periods of development can be a potential risk factor, at least in part, for developing diabetes.
'/%3e%3cuse xlink:href='%23a' transform='rotate(23.036 .093 25.536)'/%3e%3cuse xlink:href='%23a' transform='rotate(45.87 1.273 16.18)'/%3e%3cuse xlink:href='%23a' transform='rotate(69.945 .996 12.078)'/%3e%3cuse xlink:href='%23a' transform='rotate(20.66 -19.689 31.932)'/%3e%3c/svg%3e)