Chemokines are small secreted proteins functionally involved in the immune system's regulation of lymphocyte migration across numerous mammalian species. Given its growing popularity in immunological ...models, we investigated the structure and function of chemokine CXCL12 protein in tree shrews. We found that CXCL12 and its receptor CXCR4 in tree shrew had structural similarities to their homologous human proteins. Phylogenetic analysis supports the view that tree shrew is evolutionarily-close to the primates. Our results also showed that the human recombinant CXCL12 protein directly enhanced the migration of tree shrew's lymphocytes in vitro, while AMD3100 enhanced the mobilization of hematopoietic progenitor cells (HPCs) from bone marrow into peripheral blood in tree shrew in vivo. Collectively, these findings suggested that chemokines in tree shrews may play the same or similar roles as those in humans, and that the tree shrew is a viable animal model for studying human immunological diseases.
AIM: To investigate the effects of eukaryotic expression of plasmid on augmentation of liver regeneration (ALR) in rat hepatic fibrosis and to explore their mechanisms. METHODS: Ten rats were ...randomly selected from 50 Wistar rats as normal control group. The rest were administered intraperitoneally with porcine serum twice weekly. After 8 wk, they were randomly divided into:model control group, colchicine group (Col), first ALR group (ALR1), second ALR group (ALR2). Then colchicine ALR recombinant plasmid were used to treat them respectively. At the end of the 4^th wk, rats were killed. Serum indicators were detected and histopathological changes were graded. Expression of type Ⅰ, Ⅲ, collagenand TIMP-1 were detected by immunohisto-chemistry and expression of TIMP-1 mRNA was detected by semiquantified RT-PCR. RESULTS: The histologic examination showed that the degree of the rat hepatic fibrosis in two AIR groups was lower than those in model control group. Compared with model group, AIR significantly reduced the serum levels of ALT, AST, HA, LN, PCⅢ and Ⅳ (P<0.05). Immunohistochemical staining showed that expression of type Ⅰ, Ⅲ, collagenand TIMP-1 in two ALR groups was ameliorated dramatically compared with model group (I collagen: 6.94±1.42, 5.80±1.66 and 10.83±3.58 in ALR1, ALR2 and model groups, respectively; Ⅲ collagen: 7.18±1.95, 4.50±1.67 and 10.25±2.61, respectively; TIMP-1: 0.39±0.05, 0.20±0.06 and 0.53±0.12,respectively, P<0.05 or P<0.01). The expression level of TIMP-1 mRNA in the liver tissues was markedly decreased in two ALR groups compared with model group (TIMP-1 mRNA/β-actin: 0.89±0.08, 0.65±0.11 and 1.36±0.11 in ALR1, ALR2 and model groups respectively, P<0.01); CONCLUSION: ALR recombinant plasmid has beneficial effects on rat hepatic fibrosis by enhancing regeneration of injured liver cells and inhibiting TIMP-1 expressions.
Chemokines are a family of mainly-secreted proteins, traditionally associated with regulation of leukocyte trafficking during host defense and pathological immune/inflammatory reactions. All ...chemokines signal to G protein-coupled receptors. Recent studies show that chemokines and their receptors are also expressed by neuroepithelial cells, and govern developmental, physiological and pathological processes through actions towards these cells, as well as infiltrating and resident hematopoietic cells. Understanding chemokine action at the tissue level therefore requires defining which cells express chemokine receptors. At a first level of approximation (and lacking appropriate immunohistochemical reagents) this determination can be made by in situ hybridization (ISH), which localizes mRNA expression for chemokines and their receptors at the cellular level. Here we provide a protocol for ISH and demonstrate its application for localizing mRNA encoding two chemokine receptors, CXCR4 and CXCR7 in murine CNS tissues.
Stem cells reside in niches that regulate the balance between self-renewal and differentiation. The identity of a stem cell is linked with the ability to interact with its niche through adhesion ...mechanisms. To identify targets that disrupt cancer stem cell (CSC) adhesion, we performed a flow cytometry screen on patient derived glioblastoma (GBM) cells and identified junctional adhesion molecule-A (JAM-A) as a CSC adhesion mechanism essential for self-renewal and tumor growth. JAM-A was dispensable for normal neural stem/progenitor cell (NPC) function and JAM-A expression was reduced in normal brain versus GBM. Targeting JAM-A compromises the self-renewal of CSCs. JAM-A expression negatively correlated to GBM patient prognosis. Our results demonstrate that novel GBM targeting strategies can be identified through screening adhesion receptors and JAM-A represents a novel mechanism for niche driven CSC maintenance.
AIM: To directly investigate the relationship between telornerase activity and its subunit expression and the inhibitory effect of antisense hTR on pancreatic carcinogenesis.METHODS: We examined the ...telomerase activity and its subunit expression by cell culture, polymerase chain reaction(PCR), PCR-silver staining, PCR-ELISA, DNA sequencing,MTT and flow cytometry methods.RESULTS: PCR-silver staining and PCR-ELISA methods had the same specificity and sensitivity as the TRAP method.Telomerase activity was detected in the extract of the 10^th,20^th and 30^th passages of P3 cells,while it was absent in fibroblasts. Furthermore, after the 30th generation, the proliferation period of fibroblast cells was significantly prolonged. Telomerase activity and hTERTmRNA were detected in two pancreatic carcinoma cell lines, but were found to be negative in human fibroblast cells. Telomerase activity and hTERTmRNA were tested in pancreatic carcinoma specimens of 24 cases. The telomerase activity was positive in 21 of the 24 cases (87.5 %), and the hTERTmRNA in 20 cases (83.3 %). In adjacent normal tissues positive rates were both 12.5 %. There was a significant difference between the two groups. This indicated a significant correlation between the expression level of telomerase activity and histologic differentiation,metastasis and advanced clinical stage of pancreatic carcinoma. Our findings showed that the expressions of hTR and TPlmRNA were not correlated with the activity of telomerase but the expression of hTERTmRNA was. After treatment with PS-ODNs, telomerase activity in P3 cells weakened and the inhibiting effect became stronger with an increase in PS-ODNs concentration. There was a significant difference between different PS-ODN groups (P<0.05). Inhibition of telomerase activity occurred most significant with PS-ODN1.The results of the FCM test of pancreatic cancer P3 cells showed an increase in the apoptotic rate with increasing PS-ODN1 and PS-ODN2 concentrations.CONCLUSION: The expression of telomerase activity has a significant relationship to carcinogenesis. A strong correlation exists between telomerase activity and hTERTmRNA expression. The up-regulation of hTERTmRNA expression may play a critical role in human carcinogenesis.The expression of telomerase activity and its subunit level in pancreatic carcinoma significantly correlate wibh bhe dinical stage of pancreatic cardnoma and hence, may be helpful in its diagnosis and prognosis. The anti-hTR complementary to the template region of hTR is sufficient to inhibit P3 cell telomerase activity and cell proliferation in vitro, and can lead to a profound induction of programmed cell death.
To investigate the role of extracellular regulated kinase (ERK1/2) pathway in cisplatin-induced apoptosis in human ovarian carcinoma cells.
Cisplatin-induced apoptosis were stained with DAPI and was ...assessed microscopically in human epithelial adenocarcinoma ovarian cell line SKOV3 cells. ERK activation was determined by Western blotting using an anti-phospho-ERK antibody to detect ERK activity. The effect of PD98059 on ERK activity induced by cisplatin was detected by MTT assay.
Marked apoptosis of SKOV3 cells resulted from 48 hours treatment with 20 microg/mL cisplatin. Strong activation of ERK was led to by 15 microg/mL cisplatin. Dose response and time course of cisplatin induced apoptosis in SKOV3 cells. Cisplatin-induced ERK activation occurred at 12 hours and increased to highest induction at 24 hours by Western blotting. The effect of PD 98059 on ERK activity induced by cisplatin at the concentration of 100 micromol/L PD 98059. Statistically significant decreased in cell survival were observed with 100 micromol/L PD 98059 at 15 and 20 microg/mL cisplatin (P < 0.05).
Cisplatin activates the ERK signaling pathway in ovarian cancer cell line SKOV3. Inhibition of ERK activity enhances sensitivity to cisplatin cytotoxity in ovarian cancer cell line SKOV3. Evaluation of ERK activity could be useful in predicting which ovarian cancer will response most favorably to cisplatin therapy.
AIM: To construct a recombined human AFP eukaryotic expression vector for the purpose of gene therapy and target therapy of hepatocellular carcinoma (HCC),METHODS: The full length AFP-cDNA of ...prokaryotic vector was digested, and subcloned to the multi-clony sites of the eukaryotic vector. The constructed vector was confirmed by enzymes digestion and electrophoresis, and the product expressed was detected by electrochemiluminescence and immunofluorescence methods.RESULTS: The full length AFP-cDNA successfully cloned to the eukaryotic vector through electrophoresis, 0.9723 IU/ml AFP antigen was detected in the supernatant of AFPCHO by electrochemiluminescence method. Compared with the control groups, the differences were significant (P<0.05).AFP antigen molecule was observed in the plasma of AFPCHO by immunofluorescence staining. CONCLUSION: The recombined human AFP eukaryotic expression vector can express in CHO cell line. It provides experimental data for gene therapy and target therapy of hepatocellular carcinoma.
Many amphibian behaviors and physiological functions adapt to daily environmental changes through variations in circadian rhythms. However, these adaptations have yet to be reported in Dybowski's ...frog (
). We aimed to elucidate the dynamic changes in the behavior and gut microbiota of
within a 24 h cycle during their migration to hibernation sites. Thus, we monitored their behavior at 4 h intervals and collected samples for microbiome analysis. We found that the juvenile frogs arrived at hibernation sites earlier than the adults. Among the adults, the male frogs arrived earlier. The richness and diversity of the gut microbiota in the adult
were lowest at 14:00. At 6:00, the differences between the males and females were most significant. At 18:00, there was an increase in the activity of
,
,
, and
in the intestinal tracts of the male frogs, whereas in the intestinal tract of the female frogs, there was an increase in the activity of
,
,
, and
. This indicated diurnal rhythmic variations in the gut microbiota and significant sex-based differences in the microbial activity at different time points. Our findings contribute to the understanding of the circadian rhythm of
and provide crucial insights into improving breeding strategies.
Metasurfaces have attracted extensive attention in the micro/nano-optics field depending on their significant ability to modulate optical parameters. However, the current numerical simulation ...technology cannot meet the demand of the design and analysis of metasurfaces efficiently due to consuming substantial calculating time and memory. Besides, they cannot illustrate the physical mechanism straightforwardly behind the optical responses. Herein, we propose and demonstrate two equivalent circuit models systematically for a bifunctional metasurface with metal–dielectric–metal structural meta-atoms based on polarization multiplexing in the visible band. In the y-polarization state, the equivalent circuit model is established to interpret the phase shift exactly with a high goodness of fit of 0.931, resulting in the beam splitting function from the anomalous reflection phenomenon. The polychromatic light splits from 34 to 69° to produce the grating-type high-saturation structural colors with a large gamut of about 170.07% of the DCI-P3 standard. In the x-polarization state, the metasurface produces the surface lattice resonance that is suitable for the refractive index sensing function due to the high sensitivity to environmental changes. The second equivalent circuit model simulates the linewidth narrowing and red-shift phenomenon with a sensitivity relative error of 7.00%. We introduce a branch with a narrow-band-pass filter and a capacitor in series into the model to mimic the characteristic of Rayleigh anomalies. Both models extend the application of equivalent circuit theory in optics further and provide a crucial approach to enhance the insights into the mechanism of metasurfaces.
► Focus microwave system gives the stable irradiation in the enzyme immobilization. ► Micorwave irradiation accellerated the covalent immobilization of 2-deoxy-D-ribose-5-phosphate aldolase (DERA) ...and improved its activity significantly. ► The immobilized DERA under suitable microwave irradiation exihibited better thermal and storage stability.
To establish a stable and efficient immobilization technique under microwave irradiation, a focused microwave reaction system was used, where the temperature was set appropriately in the microwave system and cooling module to produce consecutive microwave irradiation. 2-Deoxy-D-ribose-5-phosphate aldolase (DERA) was rapidly and efficiently immobilized in mesocellular siliceous foams (MCFs) under microwave irradiation. When the output power in the microwave system was set to 30
W, after 3
min, 88.4% of the enzyme protein was coupled to the wall of the support pores and the specific activity of the immobilized enzyme was 2.24
U
mg
−1, 149.2% higher than that of the free enzyme and 157.0% higher than that of the non-microwave-assisted immobilized enzyme. In catalysis, microwave-assisted immobilized DERA tolerated a wider range of both pH and temperature than other DERA preparations. The thermal and storage stabilities were also significantly improved. This focused; microwave-assisted immobilization technique has proven to be simple, stable and highly efficient. This technique could also be applied to other enzyme immobilizations.
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