Stability and immunoreactivity of glycinin and β-conglycinin to hydrolysis in vitro

The present study was to compare the stability and immunoreactivity of glycinin and β-conglycinin to hydrolysis with pepsin, trypsin or cooperation of the two enzymes for different time intervals (0.5, 1, 15, 30, 60 and 120 min) at different ratios of enzyme/substrate (1:100, 1:10, 1:1 and 10:1) in vitro. The results showed that the immunoreactivity was positively related with stability of glycinin (r=0.776, P<0.05) and β-conglycinin (r=0.851, P<0.05). B polypeptide chain of glycinin was resistant to hydrolysis with trypsin, and β subunit of β-conlycinin was not liable to hydrolysis with pepsin. The two above proteins were little or not affected by incubation time and the enzyme/substrate ratio, while others' hydrolysed degree got higher with prolonging incubation time, and the hydrolysis accelerated with increasing enzyme/substrate ratio. Our results indicated digestive enzyme, incubation time and enzyme/substrate ratio had effects on stability and immunoreactivity of allergenic proteins. The most effective hydrolysis was cooperation of the two enzymes for glycinin and trypsin for β-conglycinin.     

Allergen-specific immunoglobulin,histamine and T-cell responses induced by soybean glycinin and β-conglycinin in BALB/c mice of oral sensitisation

Glycinin and β-conglycinin are major soybean allergens involved in food hypersensitivity. However, the mechanism of immune responses induced by glycinin and β-conglycinin has not been fully understood. Balb/c mice were oral sensitised with different doses (0.1, 1.0 and 10 mg/day) of soybean glycinin and β-conglycinin for five weeks. Allergen-specific immunoglobulin (Ig), serum histamine and T-cell responses were tested to assess the allergenic activity of glycinin and β-conglycinin. Mice sensitised with 0.1 or 1.0 mg/day allergens induced high levels of specific IgE, IgG1 and serum histamine compared with mice treated with saline. Furthermore, specific T-cell proliferation and significant up-regulation of interleukin (IL)-4, IL-5 and interferon (IFN)-γ were observed in splenocytes from mice gavaged with 0.1 or 1.0 mg/day soybean proteins. Low doses of glycinin or β-conglycinin can induce allergic reactions in BALB/c mice, which might be associated with increased IgE and cytokine production.     

Effects of antisense oligonucleotides on theexpression of macrophage migration inhibitoryfactor on macrophages

It is well known that the infiltration and prolifera tion of macrophages are the main features in manycases of experimental studies and human glomeru lonephritis, and they also play an important role inthe progressive renal injuries in the development ofglonerulonephritis [ 1 4 ]. Macrophage migrationfactor (MIF) is a proinflammatory cytokine thatwas originally described as a product of activatedlymphocytes that inhibited the random migration ofmacrophages in vitro and promoted the acc…     

Effects of β-carotene and aspartame on clustogenic activity of cyclophosphamide and dioxidine in mice

Antimutagenic effects of combination of aspartame (0.4 and 4 mg/kg) and -carotene (0.15-15 mg/kg) were studied by estimation of chromosome aberrations in bone marrow cells of C57Bl/6 mice. Single and 5-day treatment with this combination decreased the clastogenic effects of dioxidine and cyclophosphamide and produced a more potent and universal antimutagenic effect than its constituents.     

Effects of β-carotene and aspartame on clustogenic activity of cyclophosphamide and dioxidine in mice

Antimutagenic effects of combination of aspartame (0.4 and 4 mg/kg) and β-carotene (0.15–15 mg/kg) were studied by estimation of chromosome aberrations in bone marrow cells of C57Bl/6 mice. Single and 5-day treatment with this combination decreased the clastogenic effects of dioxidine and cyclophosphamide and produced a more potent and universal antimutagenic effect than its constituents. Translated fromByulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 130, No. 11, pp. 570–573, November, 2000     

Nitric Oxide, Superoxide, and Peroxynitrite Effects on the Insulin Secretion and Viability of βTC3 Cells

The onset of insulin-dependent diabetes mellitus (IDDM) is often associated with the infiltration of pancreatic cells by macrophages. Upon activation, macrophages release nitric oxide (NO) and superoxide (O ₂ ^- ). These species or their reactive intermediates can be cytoxic, mutagenic, or carcinogenic. Previous studies have reported both positive and negative effects of extracellularly generated NO on insulin secretion and viability of pancreatic cells. Inherent problems of several previous studies assessing the effects of NO on insulin secretion include unsteady state NO concentration exposures and the generation of other potentially damaging species. In this study, these problems were eliminated by using a modified experimental system in which NO delivery was achieved via diffusion across a gas-permeable tube and O ₂ ^- delivery was maintained using an enzymatic reaction. The delivery rates were constant, leading to steady state concentrations of NO and O ₂ ^- in the experimental system. Based on reaction kinetics, a model was developed to predict NO, O ₂ ^- and peroxynitrite ONOO^- concentrations during the experiment. This study showed that NO, O ₂ ^- and ONOO^- at predicted concentrations as high as 2.8 M, 0.25 M, and 0.1 nM, respectively, do not affect the insulin secretion rates of TC3 pancreatic cells over short times. © 2000 Biomedical Engineering Society. PAC00: 8717-d     

Effects of age and β-amyloid on cognitive changes in normal elderly people

Age-related decline is common in multiple cognitive domains. β-amyloid (Aβ) deposition, a pathological hallmark of Alzheimer's disease, is also associated with cognitive changes in many older people. In this study, we examined a wide range of cognitive function in order to differentiate the effect of age and Aβ on cognition during aging. Using positron emission tomography (PET) imaging with the radiotracer Pittsburgh Compound B (PIB), we classified normal older subjects as High PIB-Old and Low PIB-Old and applied sequential multivariate analyses (i.e., principal components analysis [PCA] and discriminant analysis) to obtain summary measures of cognitive tests encompassing multiple cognitive domains. Among 5 cognitive components, a significant age effect was observed in component scores of visual memory and executive functions, regardless of the level of Aβ. Discriminant scores (weighted scores of the 5 cognitive components) revealed a significant effect of both age and Aβ and were further associated with quantitative PIB counts. The results of the current study highlight both effects of age and Aβ on cognitive changes in normal elderly.     

Effects of clenbuterol and ICI118551, a selective β2-antagonist,on the growth of skeletal muscle of suckling rats

The ₂-adrenergic agonist, clenbuterol, was administered to lactating rats (4 mg/kg diet) from post-partum day 1 to day 19, or directly injected into neonate rats (0.1 and 1.0 mg/kg body weight) from post-partum day 3 until day 15. Changes in body weight and the skeletal muscles soleus (SOL) and extensor digitorum longus (EDL) were studied in both dams and suckling offspring. Drug treatment consistently increased body weight in dams whilst significantly reducing the growth of their suckling pups. In dams treated with clenbuterol (4 mg/kg of diet) muscle weights and protein contents were significantly increased. Total protein content increased by 16% in SOL and 47% in EDL after 19 days of treatment. In contrast, in their suckling pups, there was a 22% and 26% reduction in protein content of SOL and EDL respectively. Administration of the ₂-antagonist ICI1 18551 to these pups failed to prevent these reductions in body and muscle weights. Hence, if clenbuterol did reach the pups via the milk from treated mothers it did not act via conventional ₂-receptors. Injection of pups with clenbuterol (1.0 mg/kg every 12 h) from litters suckling from untreated dams also resulted in significant reductions in muscle weights and protein contents. Protein content was reduced by 10% in SOL and 13% in EDL after 12 days of treatment. No alteration in fibre type proportions in SOL or EDL resulted from this treatment. Further work is required to determine whether the growth suppression in the two situations occurs via the same mechanism.     

Nuclear β-catenin is increased in systemic sclerosis pulmonary fibrosis and promotes lung fibroblast migration and proliferation

Pulmonary fibrosis is a disease that results in loss of normal lung architecture, but the signaling events that drive tissue destruction are incompletely understood. Wnt/β-catenin signaling is important in normal lung development, but whether abnormal signaling occurs in lung fibrosis due to systemic sclerosis and the consequences of β-catenin signaling toward the fibrogenic phenotype remain poorly defined. In this study, we show nuclear β-catenin accumulation in fibroblastic foci from lungs of patients with systemic sclerosis-associated advanced pulmonary fibrosis. Forced activation of β-catenin signaling in three independently derived sources of normal human lung fibroblasts promotes proliferation and migratory activities but is not sufficient to activate classic markers of fibroblast activation, such as TGF-β, type 1 collagen, α-smooth muscle actin, and connective tissue growth factor. These findings indicate that activation of β-catenin signaling in pulmonary fibroblasts may be a common feature of lung fibrosis, contributing to fibroproliferative and migratory activities associated with the disease.     

Effects of Fas,NF-κB and caspases on rat microvascular endothelial cell apoptosis induced by TNFα

Effects of Fas,NF-κB and caspases on rat microvascular endothelial cell apoptosis induced by TNFα     

Atorvastatin effects on proliferation and apoptosis of rat bone marrow-derived endothelial progenitor cells in vitro北大核心

BACKGROUND: Atorvastatin has a cardiovascular protective effect that significantly improves endothelial function and promotes the mobilization, migration, and differentiation of endothelial progenitor cells. However, the screening of atorvastatin concentration for in vitro cell culture is not well documented. OBJECTIVE: To investigate the effects of different concentrations of atorvastatin on rat bone marrow-derived EPCs growth characteristics. METHODS: Bone marrow mononuclear cells from Sprague-Dawley rats were induced in selective culture fluid to culture EPCs. Immunofluorescence staining was used to identify cell surface markers. Harvested EPCs were divided into control group and atorvastatin groups with four different concentrations (0.01, 0.1, 1, and 10 µmol/L) for culture. The growth and proliferation of EPCs were observed under light microscope and MTT assay. Flow cytometry was used to detect apoptosis in EPCs. Nitric oxide and endothelial nitric oxide synthase levels in the culture fluid were measured by nitrate reductase method. RESULTS AND CONCLUSION: The number of cells tended to increase in the control and atorvastatin groups, and it was highest in the 1 µmol/L atorvastatin group. The cell number in the 10 µmol/L atorvastatin group began to decrease at 7 days of culture. Among the five groups, the apoptotic rate of cells was lowest in the 1 µmol/L atorvastatin group and highest in the 10 µmol/L atorvastatin group. The levels of nitric oxide and endothelial nitric oxide synthase were significantly higher in the 0.01, 0.1 and 1.0 µmol/L atorvastatin groups compared with the control group (P < 0.01), but lower in the 10 µmol/L atorvastatin group compare with the other groups (P < 0.01). Overall, atorvastatin can promote the proliferation of endothelial progenitor cells and reduce apoptosis by increasing the production of endothelial nitric oxide synthase and nitric oxide, and 1 µmol/L atorvastatin is most suitable for the EPCs culture. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Effects of tumour necrosis factor α and interleukin 1 β on the proliferation of cultured glomerular epithelial cells

Rat glomerular epithelial cells were cultured with human monocyte supernatant or with recombinant cytokines. A primary glomerular culture and a glomerular epithelial cell culture were made; supernatant from monocyte cultures derived from healthy humans, and recombinant tumour necrosis factor (TNF ) or recombinant interleukin 1 (IL-1 ) were added. Cell proliferation rates were assayed by the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay. In serum-free media, consistent proliferation of glomerular epithelial cells (GEC) was observed throughout the 3 week culture period. Significant growth-stimulatory effects were induced by lipopolysaccharide-treated monocyte conditioned medium and by 1–50 ng/ml of TNF , growth being up to 400% more than in the control culture. The effect of TNF depended mainly on its interaction with epidermal growth factor (EGF). In contrast to TNF , IL-1 inhibited GEC proliferation; this was due to the early appearance and proliferation of mesangial cells, despite the culture being serum-free. This study showed that activated monocytes secrete growth factors for GEC in vitro, and that interaction between both TNF and IL-1 and between TNF and EGF can modulate GEC proliferation. These findings suggest that, under pathological conditions, monocytes or macrophages affect GEC proliferation, probably being involved in crescent formation.     

Effects of Hydrocortisone and β-Aminopropionitrile on Stress-Strain and Stress-Relaxation Behaviors,and Birefringent Retardation of Collagen Fibers in the Rat Incisor Periodontal Ligament

Three groups of male Wistar rats received daily subcutaneous injections of 10 mg/kg of hydrocortisone (HC group), 300 mg/kg of &#103 -aminopropionitrile (BAPN group), or saline (control group), for 10 days. The shear stress-strain and stress-relaxation properties of the incisor periodontal ligament were examined in transverse sections from dissected mandibles. Both the maximum shear stress and failure strain energy density increased significantly following the administration of hydrocortisone. The maximum shear stress decreased following the administration of BAPN. However, the stress-relaxations in the initial 10 min did not show significant differences among the three groups. Polarized light microscopic analysis revealed that the retardation value of the collagen fibers was highest in the HC group and lowest in the BAPN group for the bone-related area, but not for the tooth-related and middle areas of the ligament. It is suggested that the changes induced by hydrocortisone or BAPN occurred mainly in the elastic components and to a minor extent in the viscous components although the physical and biomechanical properties are determined by the interaction of all the various components. We also suggest that the main response to the drugs occurred in the collagen fibers in the bone-related area of the ligament.     

Effects of different amyloid β-protein analogues on synaptic function

Perisynaptic accumulations of amyloid β-protein (Aβ) play a critical role in the synaptic dysfunction underlying the cognitive impairment observed in Alzheimer’s disease. The methionine residue at position 35 (Met35) in Aβ is highly subject to oxidation in Alzheimer’s disease brains. In hippocampal brain slices we found that long-term potentiation at CA3–CA1 synapses was significantly inhibited by wild type Aβ42 in which Met35 is reduced, but not by Aβ42 harboring Met35 sulfoxide. Similar differences were observed when basal synaptic transmission was investigated in autaptic hippocampal neurons. The significant decreases in excitatory postsynaptic current amplitude, vesicle release probability and miniature excitatory postsynaptic current frequency caused by 20-minute exposure to wild type Aβ42 were not observed after exposure to Aβ42 harboring Met35 sulfoxide. With longer (24-hour) Aβ treatments, this early impairment of the presynaptic terminal function extended to involve the postsynaptic side as well. The Met35 oxidation also affected Aβ42 negative impact on dendritic spine density and expression of pre- and postsynaptic proteins (synaptophysin and postsynaptic density protein-95). Our findings suggest that oxidation of Met35 is critical for molecular, structural, and functional determinants of Aβ42 synaptotoxicity.     

Effects of synthetic bis-β-chloroethylamine estrogen derivatives on proliferation of skin sarcoma cells

The effects of four new synthetic bis-β-chloroethylamine-containing estrogens and known cytostatic agents chlorophenacyl and estradiol mustard were compared on monolayer cultures of transformed L-929 fibroblasts (from murine skin sarcoma). The drugs within the concentration range of 10⁻⁵-5×10⁻⁷M inhibited proliferation of cultured cells by 67%. Chlorophenacyl displayed the least antiproliferative activity (15% inhibition at 10⁻⁵M). Steroid nucleus introduced into the molecule enhanced antiproliferative activity of test drug in comparison with chlorophenacyl, probably due to accumulation of the hormone-cytostatic molecules in cells. Estradiol had no effect on proliferative activity of L-929 cells, and no specific estrogen-binding sites were found in cultured transformed fibroblasts. The antiproliferative effect of hormone-cytostatics on this culture is not mediated via specific interactions with estrogen receptors. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 129, No. 6, pp. 695–697, June, 2000     

Effects of β-escin and saponin on the transverse-tubular system and sarcoplasmic reticulum membranes of rat and toad skeletal muscle

 Mechanically skinned skeletal muscle fibres from rat and toad were exposed to the permeabilizing agents β-escin and saponin. The effects of these agents on the sealed transverse tubular system (t-system) and sarcoplasmic reticulum (SR) were examined by looking at changes in the magnitude of the force responses to t-system depolarization, the time course of the fluorescence of fura-2 trapped in the sealed t-system, and changes in the magnitude of caffeine-induced contractures following SR loading with Ca²⁺ under defined conditions. In the presence of 2 μg ml^–1β-escin and saponin, the response to t-system depolarization was not completely abolished, decreasing to a plateau, and a large proportion of fura-2 remained in the sealed t-system. At 10 μg ml^–1, both agents abolished the ability of both rat and toad preparations to respond to t-system depolarization after 3 min of exposure, but a significant amount of fura-2 remained in sealed t-tubules even after exposure to 100 μg ml^–1β-escin and saponin for 10 min. β-Escin took longer than saponin to reduce the t-system depolarizations and fura-2 content of the sealed t-system to a similar level. The ability of the SR to load Ca²⁺ was reduced to a lower level after treatment with β-escin than saponin. This direct effect on the SR occurred at much lower concentrations for rat (2 μg ml^–1β-escin and 10 μg ml^–1 saponin) than toad (10 μg ml^–1β-escin and 150 μg ml^–1 saponin). The reverse order in sensitivities to β-escin and saponin of t-system and SR membranes indicates that the mechanisms of action of β-escin and saponin are different in the two types of membrane. In conclusion, this study shows that: (1) β-escin has a milder action on the surface membrane than saponin; (2) β-escin is a more potent modifier of SR function; (3) simple permeabilization of membranes is not sufficient to explain the effects of β-escin and saponin on muscle membranes; and (4) the t-system network within muscle fibres is not a homogeneous compartment. Received: 18 November 1998 / Received after revision: 6 January 1999 / Accepted: 7 January 1999     

Fentanyl inhibits proliferation and invasion of colorectal cancer via β-catenin

Background and aim: Fentanyl is widely used for relieving pain and narcotizing in cancer patients. However, there are few published reports regarding the effects of fentanyl on tumor control and treatment. Here we investigated the effects of fentanyl on tumor growth and cell invasion in the human colorectal carcinoma (HCT116) cells. Methods: Nude mice xenografts of HCT116 cells were established to assess the inhibition effect on tumor growth by fentanyl. MTT and Transwell were employed to determine the cell survival rate and cell invasion, respectively. MicroRNAs and mRNAs expression were quantified by real-time PCR. β-catenin and matrix metalloproteinases (MMP-2 and MMP-9) expression were assayed by western blotting. β-Catenin-specific small interfering RNA (Si-β-catenin) and miR-182 mimics were transfected in cells to investigate the mechanism underlying the effects of fentanyl on the colorectal tumor and HCT116 cells. Results: Treatment with fentanyl inhibited the tumor growth and HCT116 cells invasion. Fentanyl also downregulated the expression of β-catenin and miR-182 in both xenograft tumors and HCT116 cells, and decreased the protein level of MMP-9 in HCT116 cells. Downregulation of β-Catenin resulted in the decrease of miR-182 expression in colorectal cells. In addition, the overexpression of miR-182 reversed the effect of fentanyl on MMP-9 expression and cell invasion of HCT116 cells. Conclusions: The current study demonstrated that the inhibition of tumor growth and cell invasion in colorectal cancer by fentanyl is probably due to downregulation of miR-182 and MMP-9 expression by β-catenin.     

Comparative study on the stability of soybean (Glycine max) β-conglycinin in vivo

The immunoreactivity and structural variation of β-conglycinin in digesta from digestive tracts of pigs were measured by inhibition ELISA and sodium dodecyl sulphate polyacrylamide gel electrophoresis, respectively. Results showed that the immunoreactivity disappearance proportion of β-conglycinin significantly increased from stomach to the caecum in all groups (P<0.05). In the stomach, upper-jejunum and middle-jejunum, the immunoreactivity disappearance proportion of β-conglycinin significantly increased among these three groups (P<0.05), while it has no significant difference in ileum and caecum (P>0.05). The α′, α subunits of β-conglycinin were easier to digest than the β subunit. It indicated that immunoreactivity disappearance proportion of β-conglycinin tended to increase with the growth of age and the descending down of digestive tract, while the β subunits of β-conglycinin are more stable to digestion than α′, α subunits.     

Pyrrolidine dithiocarbamate inhibits nuclear factor-κB pathway activation, and regulates adhesion, migration, invasion and apoptosis of endometriotic stromal cells

The activation of nuclear factor-κB (NF-κB) has been implicated in the development and progression of endometriosis. The aim of this study is to investigate the potential application of pyrrolidine dithiocarbamate (PDTC), a potent NF-κB inhibitor, in the treatment of endometriosis. NF-κB-DNA-binding activity, IκB phosphorylation and expression of nuclear p65 protein in endometriotic ectopic stromal cells (EcSCs), endometriotic eutopic stromal cells (EuSCs) and normal endometrial stromal cells (NESCs) were detected by electrophoretic mobility shift assay and western blot analysis. Adhesion, migration, invasion and apoptosis of EcSCs were observed by means of adhesion, migration, invasion and terminal deoxynucleotidyl transferase-mediated dUDP nick-end labeling assay, respectively. Gene and protein expressions of CD44s, matrix metalloproteinase (MMP)-2, MMP-9 and survivin in EcSCs were measured by RT-PCR and western blot analysis. The results showed that PDTC in the absence or presence of interleukin (IL)-1β showed stronger inhibitory effects on NF-κB-DNA-binding activity, IκB phosphorylation and expression of nuclear p65 protein in EcSCs than those in EuSCs or NESCs. PDTC enhanced apoptosis, and suppressed IL-1β-induced cellular adhesion, migration and invasion of EcSCs. Pretreatment of EcSCs with PDTC attenuated IL-1β-induced expressions of CD44s, MMP-2, MMP-9 and survivin at gene and protein levels. All these findings suggest that PDTC induces apoptosis and down-regulates adhesion, migration and invasion of EcSCs through the suppression of various molecules. Therefore, PDTC could be used as a therapeutic agent for the treatment of endometriosis.