[The effect of curcumin on the expression of miR-155 to apoptosis and invasion of extravillus trophoblast cells treated by lipopolysaccharide].

Objective: To investigate the effect of curcumin on the expression of miR-155, apoptosis and invasion of extravillus trophoblast cells treated by lipopolysaccharide (LPS). Methods: Human trophoblast cells (HTR-8/SVneo cells) were divided into 4 groups, the curcumin + LPS group (pre-treated by curcumin of 12.5, 25, 50 µmol/L, then LPS of 100 ng/ml), the LPS group (100 ng/ml), the recombinant adenovirus group (miR-155, multiplicity of infection100) and the control group. The miR-155 level in HTR-8/SVneo cells was measured by real-time PCR, and the expression of NF-κB was analyzed by luciferase gene expression. The apoptosis of HTR-8/SVneo cells was tested by cell death detection ELISA and the level of NF-κB in HTR-8/SVneo cells was measured by western blot. In addition, transwell was used to test the invasive ability of HTR-8/SVneo cells in all the groups. Results: (1) The intracellular expression of miR-155 in the LPS group was (2.13±0.22) times of the control group (P<0.01); and the expressions of miR-155 in 12.5, 25, 50 µmol/L curcumin+LPS groups were (0.37±0.08) , (0.68±0.14) , (0.49±0.09) times as the LPS group, with statistically significant difference (P<0.05). (2) The expreesion of NF-κB in the LPS group was (2.25±0.56) times of the control group. The expreesion of NF-κB in the 12.5, 25, 50 µmol/L curcumin+LPS groups were (0.80±0.07) , (0.74±0.05) , (0.49±0.19) times to the LPS group, with statistically significant difference(P<0.01). (3)The p65 protein in the LPS group was (1.50±0.22) , significantly higher than the control group (0.95±0.25, P<0.01) . In 12.5, 25, 50 µmol/L curcumin+LPS groups, the p65 protein were (0.31±0.07) , (0.75±0.14) , (0.49±0.08) . Compared with the LPS group, p65 was down-regulated by curcumin, with statistically significant difference (P<0.05). (4) In the cell death detection ELISA, the A value in the control group, the LPS group and the recombined adenovirus group were (0.89±0.17) , (2.02±0.35) and (1.67±0.48) , respectively, with statistically significant difference (P<0.05). In the curcumin+LPS groups, when the curcumin concentrations were 25 or 50 µmol/L, the A value were (0.74±0.05) , (0.49±0.09) , respectively, compared with the LPS group(set as 1.00), with statistically significant difference (P<0.05). When the curcumin concentration was 12.5 µmol/L, the A value was (0.80±0.07) , with no statistical significance (P>0.05). (5) The transmembrane cells were (47±8), (60±14) in the LPS group and the recombined adenovirus group, respectively. Compared with the control group (169±18), the differences were statistically significant (P<0.05). In the curcumin + LPS groups, the transmembrane cells were (143±10), (137±10) when the curcumin concentrations were 12.5, 25 µmol/L, significantly higher than the LPS group (P<0.05) . The transmembrane cells were (55±7) when the curcumin concentration was 50 µmol/L, with no statistical difference(P>0.05). Conclusion: The treatment of curcumin could downregulate the expression of NF-κB/miR-155, thus inhibit NF-κB signal pathway and the apoptosis of extravillus trophoblast cells, and protect their invasive ability.

BMP-2 promotes chondrogenesis of rat adipose-derived stem cells by using a lentiviral system.

Osteoporosis poses a major public health threat in aging societies. Adipose-derived stem cells (ADSCs) are multipotent adult stem cells that have the ability to yield mesenchymal stem cells, and have the potential to undergo osteogenesis and bone regeneration. Bone morphogenetic proteins (BMPs) have been demonstrated to upregulate bone gene expression after mechanical injury and to improve bone injury repair. This study aimed to produce BMP-2 expression in ADSCs by using lentiviral vectors. Subcutaneous adipose tissue from 4-week-old male Sprague-Dawley rats was used. Oil red O staining was used to detect adipocyte formation from ADSCs. Induction of ADSC osteogenesis was confirmed with Alizarin red S staining. The recombinant lenti-hBMP-2/neo was constructed to infect ADSCs, BMP-2 expression was measured by immunoblotting analysis, and cellular alkaline phosphatase levels were examined. We found that >70% of ADSC cells could be induced to differentiate into osteocytes or adipocytes. Under osteogenic induction, ADSCs showed increased intracellular calcium deposition, the formation of calcium tubercles, and the disappearance of cellular structures in calcium tubercles. After infection of ADSCs by lenti-hBMP-2/neo, BMP-2 was expressed after doxycycline induction. We, thus, conclude that ADSCs maintain vigorous growth ex vivo and possess stem cell-like properties. When infected with lenti-hBMP-2/neo, ADSCs can be induced to promote BMP-2 expression.

[Expression, characterization of recombinant human soluble BAFF secreted from CHO cell].

B cell-activating factor of the TNF family (BAFF) is critical for B cell maturation and survival. Here, we constructed a stable CHO cell line, in which the expression level of soluble form of BAFF (sBAFF) was raised from 0.13 microg/ml to 0.55 microg/ml. Purified recombinant sBAFF from these CHO cells not only bound to its receptors but also co-stimulated the proliferation of human peripheral blood B lymphocyte in vitro. These results provided us with a useful basis for further studies about sBAFF-related research.

[Detection of Plasmodium falciparum using the plasmid Pf rep 20 probe (preliminary report)].

A cloned repetitive DNA sequence (rep 20) labeled with 32P was evaluated as diagnostic probe for P. falciparum in 39 blood samples from Hainan and Yunan provinces. Its specificity and sensitivity were studied and compared with total genomic DNA probe isolated from P. f. Hainan Fcc1/HN isolate. The pPF rep 20 probe recognized the DNA of P. f. Hainan, Yunnan and Anhui isolates, but did not hybridize with the DNA of P. vivax, P. cynomolgi, P. knowlesi and P. yoelii. The genomic probe hybridized with the DNA of 3 isolates and cross-hybridized with the DNA of all the other Plasmodium species tested, but did not hybridize with host DNA. The plasmid rep 20 probe was able to detect parasitemia level of 0.001% in 20 microliters blood from culture and 10pg DNA of 3 P. f. isolates after 3 days film exposure. It could hybridize with the blood samples of P. f. patients from Hainan and Yunnan with a sensitivity of 95% (37/39). The genomic probe could detect the same parasitemia and DNA levels as plasmid rep 20 probe for Yunnan and Anhui isolates, and 0.0001% parasitemia and 1 pg DNA for Hainan isolate. It had a sensitivity of 97% (38/39) when used to detect patient samples. The results indicated that plasmid rep 20 probe was specific and rather sensitive to DNA of P. falciparum isolates from China and may be useful in epidemiological studies.