[Effects of bone marrow mesenchymal stem cell transplantation on retinal neovascularization in neonatal rats with oxygen-induced retinopathy].

OBJECTIVE: To explore the effects of rat bone mesenchymal stem cell (BMSC) transplantation on retinal neovascularization, and to observe the changes of hypoxia-inducible factor-1 alpha (HIF-1α) and vascular endothelial growth factors (VEGF) in rats with oxygen-induced retinopathy (OIR). METHODS: Seventy-two seven-day-old Sprague-Dawley rats were randomly divided into three groups: normal control (CON), model (OIR) and BMSC transplantation. In the BMSC transplantation group, BMSCs were transplanted 5 days after oxygen conditioning. The phosphate buffered saline of the same volume was injected in the CON and OIR groups. The OIR model was prerpared according to the classic hyperoxygen method. At seven days after transplantation, retinal neovascularization was examined by retinal flat-mount staining and hematoxylin eosin (HE) staining. The expression of HIF-1α and VEGF proteins was examined by immunohistochemistry staining and Western blot analysis. RESULTS: The retinal flat-mount staining results showed that the vessels were well organized in the CON group, but the vessels were irregularly organized, and lots of nonperfusion areas were observed in the OIR group. The large vessels were a bit circuitous, the retinal vessels were relatively organized, and less nonperfusion areas were noted in the BMSC transplantation group. The HE staining results showed that many neovessels and preretinal neovascular (pre-RNC) cells were observed on the internal limiting membrane in the OIR group. There were less pre-RNC cells in the BMSC transplantation group compared with the OIR group (P<0.01). The immunohistochemistry analysis showed that more HIF-1α+ and VEGF+ cells were observed in the OIR group compared with the CON group, and less HIF-1α+ and VEGF+ cells were observed in the BMSC transplantation group compared with OIR group (P<0.05). The Western blot analysis showed the expression of HIF-1α and VEGF proteins in the OIR group was significantly higher than that in the CON group. The expression of HIF-1α and VEGF proteins in the BMSC transplantation group was lower than that in the OIR group (P<0.01). CONCLUSIONS: BMSC transplantation therapy could alleviate retinal neovascularization in OIR rats, and its mechanisms might be associated with the inhibition of the expression of HIF-1α and VEGF proteins.

Comparative characterization of recombinant ZZ protein-alkaline phosphatase and its application in enzyme immunoassays.

A functional fusion protein, which consists of an antibody and an enzyme that can be used in enzyme immunoassays, has been constructed. However, a quantitative comparison of the characteristics of fusion proteins and chemical conjugates of the parents, which are functionally produced in a uniform microbial system, has not been adequately achieved. In this study, a fusion protein between the ZZ protein and Escherichia coli alkaline phosphatase (AP) and the parental ZZ protein and AP for chemical conjugate was functionally produced in the same bacterial system. A detailed examination of the ZZ-AP fusion protein and the effect of the ZZ-AP chemical conjugate on IgG affinity and enzymatic activity were performed. Compared with the parents, the equilibrium dissociation constant of ZZ-AP conjugate decreased by 32 % and catalytic activity decreased by 24 %, whereas the ZZ-AP fusion retained full parental activities and exhibited an approximately tenfold higher sensitivity than that of ZZ-AP conjugate in enzyme-linked immunosorbent assay. Thus, ZZ-AP fusion is a promising immunoreagent for IgG detection and a potential biolinker between antibodies and reporter enzymes (i.e., IgG-ZZ-AP fusion complex) in immunoassays.

[Effects of umbilical cord blood monocytes transplantation on EPO protein and oligodendrocyte progenitors in neonatal rats with hypoxic-ischemic brain damage].

OBJECTIVE: To study the effects of umbilical cord blood monocytes (UCBMC) transplantation on erythropoietin (EPO) protein and oligodendrocyte progenitor cells in hypoxia-ischemia (HI) neonatal rats. METHODS: Forty seven-day-old Sprague-Dawley rats were randomly divided into normal control (N), HI, UCBMC and HI+UCBMC groups (n=10 each). Hypoxic-ischemic brain damage (HIBD) model was prepared according to the Rice method. Twenty-four hours after hypoxia, the N and HI groups were injected with 2 µL phosphate buffered saline (PBS), and the UCBMC and HI+UCBMC groups were injected with 3×10(6) UCBMC via the lateral ventricle. EPO protein and oligodendrocyte progenitor cells in the subventricular zone of the injured brain were observed by EPO/DAPI and NG2/DAPI immunofluorescence double staining, and their correlation was analyzed. RESULTS: Seven days after transplantation, there were more NG2(+)DAPI(+) and EPO(+)DAPI(+) cells in the HI+UCBMC group than in the UCBMC (P<0.05), N and HI groups (P<0.01). More NG2(+)DAPI(+) and EPO(+)DAPI(+) cells were observed in the UCBMC group compared with the N and HI groups (P<0.01). There were more NG2(+)DAPI(+) cells in the N group than in the HI group (P<0.01). The number of NG2(+)DAPI(+) cells was correlated with the number of EPO(+)DAPI(+) cells in the HI+UCBMC group (r=0.898, ß=1.4604, P<0.01). CONCLUSIONS: UCBMC can promote expression of oligodendrocyte progenitor cells, which is correlated with an increase in EPO protein and thus repairs brain white matter damage in neonatal rats with HIBD.

[Effects of bone marrow mesenchymal stem cell transplantation on retinal cell apoptosis in premature rats with retinopathy].

OBJECTIVE: To explore the effects of marrow mesenchymal stem cell (BMSC) transplantation on retinal cells apoptosis and changes to neurotrophin-3 (NT-3 and ciliary neurotrophic factor (CNTF) in rats with retinopathy of prematurity (ROP). METHODS: Seven-day-old Sprague-Dawley rats were randomly divided into normal control (CON), ROP, BMSC transplantation (BMSCs were transplanted 5 days after oxygen conditioning) and phosphate buffered saline (PBS) groups. The ROP model was prepared according to the classic hyperoxygen method. Seven days after transplantation, TUNEL/DAPI, NT-3/API and CNTF/DAPI double-labeled immunofluorescence were used to examine the effects of BMSC transplantation on both the apoptosis of retinal cells and the expression of NT-3 and CNTF protein in the retinal cells of the ROP rats. RESULTS: Seven days after BMSC transplantation, there were few TUNEL+ DAPI+ cells observed in the CON group. There were fewer TUNEL+DAPI+ cells observed in the BMSC group than in the ROP group (P<0.01), but there was no significant difference between the ROP and PBS groups (P>0.05). There were few NT-3+DAPI+ cells and CNTF+DAPI+ cells in the CON group. There were more NT-3+DAPI+ and CNTF+DAPI+ cells in the ROP group than in the CON group, but there was no significant difference between the ROP and CON groups (P>0.05). More NT-3+DAPI+ and CNTF+DAPI+ cells were observed in the BMSC group compared with the ROP group (P<0.01), and there was no significant difference in either NT-3+DAPI+ or CNTF+DAPI+ cells between the ROP and PBS groups (P>0.05). CONCLUSIONS: BMSC transplantation therapy could alleviate the apoptosis of retinal cells in ROP rats, and its mechanisms might be associated with promoting the expression of NT-3 and CNTF protein in retinal cells.

Chromodomain Helicase/ATPase DNA-Binding Protein 1-Like Gene (CHD1L) Expression and Implications for Invasion and Metastasis of Breast Cancer.

BACKGROUND: Chromodomain helicase/ATPase DNA-binding protein 1-like gene (CHD1L), also known as ALC1 (amplified in liver cancer 1 gene), is a new oncogene amplified in many solid tumors. Whether this gene plays a role in invasion and metastasis of breast cancer is unknown. METHODS: Immunohistochemistry was performed to detect the expression of CHD1L in patients with invasive ductal carcinoma and normal mammary glands. Chemotaxis, wound healing, and Transwell invasion assays were also performed to examine cell migration and invasion. Western blot analysis was conducted to detect the expression of CHD1L, MMP-2, MMP-9, pAkt/Akt, pARK5/ARK5, and pmTOR/mTOR. Moreover, ELISA was carried out to detect the expression levels of MMP-2 and MMP-9. Nude mice xenograft model was used to detect the invasion and metastasis of breast cancer cell lines. RESULTS: CHD1L overexpression was observed in 112 of 268 patients (41.8%). This overexpression was associated with lymph node metastasis (P = 0.008), tumor differentiation (P = 0.020), distant metastasis (P = 0.026), MMP-2 (P = 0.035), and MMP-9 expression (P = 0.022). In the cell experiment, reduction of CHD1L inhibited the invasion and metastasis of breast cancer cells by mediating MMP-2 and MMP-9 expression. CHD1L knockdown via siRNA suppressed EGF-induced pAkt, pARK5, and pmTOR. This knockdown inhibited the metastasis of breast cancer cells into the lungs of SCID mice. CONCLUSIONS: CHD1L promoted the invasion and metastasis of breast cancer cells via the PI3K/Akt/ARK5/mTOR/MMP signaling pathway. This study identified CHD1L as a potential anti-metastasis target for therapeutic intervention in breast cancer.