Hydrogen Repairs LPS-Induced Endothelial Progenitor Cells Injury via PI3K/AKT/eNOS Pathway.

Endotoxins and other harmful substances may cause an increase in permeability in endothelial cells (ECs) monolayers, as well as ECs shrinkage and death to induce lung damage. Lipopolysaccharide (LPS) can impair endothelial progenitor cells (EPCs) functions, including proliferation, migration, and tube formation. EPCs can migrate to the damaged area, differentiate into ECs, and participate in vascular repair, which improves pulmonary capillary endothelial dysfunction and maintains the integrity of the endothelial barrier. Hydrogen (H2) contributes to the repairment of lung injury and the damage of ECs. We therefore speculate that H2 protects the EPCs against LPS-induced damage, and it's mechanism will be explored. The bone marrow-derived EPCs from ICR Mice were treated with LPS to establish a damaged model. Then EPCs were incubated with H2, and treated with PI3K inhibitor LY294002 and endothelial nitric oxide synthase (eNOS) inhibitor L-NAME. MTT assay, transwell assay and tube formation assay were used to detect the proliferation, migration and angiogenesis of EPCs. The expression levels of target proteins were detected by Western blot. Results found that H2 repaired EPCs proliferation, migration and tube formation functions damaged by LPS. LY294002 and L-NAME significantly inhibited the repaired effect of H2 on LPS-induced dysfunctions of EPCs. H2 also restored levels of phosphor-AKT (p-AKT), eNOS and phosphor-eNOS (p-eNOS) suppressed by LPS. LY294002 significantly inhibited the increase of p-AKT and eNOS and p-eNOS expression exposed by H2. L-NAME significantly inhibited the increase of eNOS and p-eNOS expression induced by H2. H2 repairs the dysfunctions of EPCs induced by LPS, which is mediated by PI3K/AKT/eNOS signaling pathway.

Linc00426 accelerates lung adenocarcinoma progression by regulating miR-455-5p as a molecular sponge.

Increasing lines of evidence indicate the role of long non-coding RNAs (LncRNAs) in gene regulation and tumor development. Hence, it is important to elucidate the mechanisms of LncRNAs underlying the proliferation, metastasis, and invasion of lung adenocarcinoma (LUAD). We employed microarrays to screen LncRNAs in LUAD tissues with and without lymph node metastasis and revealed their effects on LUAD. Among them, Linc00426 was selected for further exploration in its expression, the biological significance, and the underlying molecular mechanisms. Linc00426 exhibits ectopic expression in LUAD tissues and cells. The ectopic expression has been clinically linked to tumor size, lymphatic metastasis, and tumor differentiation of patients with LUAD. The deregulation of Linc00426 contributes to a notable impairment in proliferation, invasion, metastasis, and epithelial-mesenchymal transition (EMT) in vitro and in vivo. Mechanistically, the deregulation of Linc00426 could reduce cytoskeleton rearrangement and matrix metalloproteinase expression. Meanwhile, decreasing the level of Linc00426 or increasing miR-455-5p could down-regulate the level of UBE2V1. Thus, Linc00426 may act as a competing endogenous RNA (ceRNA) to abate miR-455-5p-dependent UBE2V1 reduction. We conclude that Linc00426 accelerates LUAD progression by acting as a molecular sponge to regulate miR-455-5p, and may be a potential novel tumor marker for LUAD.

[MiR-204 inhibits invasion and metastasis of breast cancer cells by targeted regulation of HNRNPA2B1].

OBJECTIVE: To investigate the effect of miR-204 on the invasion and metastasis of breast cancer by targeted regulation of HNRNPA2B1. METHODS: The bioinformatics database was used to obtain data of the expressions of miR-204 in breast cancer patients and the survival rate of the patients. RT-qPCR was used to detect the expression of miR-204 in breast cancer cell lines. The expression vector GV369-miR-204 was used to overexpress miR-204 in MDA-MB-231 cells. Transwell assay was performed to detect the effect of miR-204 on the migration and invasion ability of the breast cancer cells. The key genes (hub genes) of miR-204 were determined by bioinformatics method. A dual luciferase assay was used to analyze the targeting relationship between miR-204 and HNRNPA2B1. The expression of HNRNPA2B1 in MDA-MB-231 cells after miR-204 overexpression was detected by Western blotting, and Transwell assay was used to examine the changes in the cell invasion ability. RESULTS: The expression of miR-204 was decreased in both breast cancer tissues, and was significantly lower in breast cancer MDA-MB-231 cells than in MCF-10A cells (P < 0.05). The decreased expression of miR-204 was associated with poorer prognosis of breast cancer patients (P < 0.05). Upregulation of miR-204 in MDA-MB-231 cells significantly inhibited the invasion and migration of the cells (P < 0.05). Analysis of the data from the Starbase revealed that the expression of miR-204-5p was negatively correlated with the expression of HNRNPA2B1, and the expression of HNRNPA2B1 was increased in breast cancer patients (P < 0.05) in association with a poorer prognosis of the patients (P < 0.05). Dual luciferase assay demonstrated that miR-204 could bind to HNRNPA2B1 in a target-specific manner. Western blotting and Transwell assay showed that miR-204 significant inhibited the migration and invasion ability of breast cancer cells by targeting HNRNPA2B1 (P < 0.05). CONCLUSIONS: miR-204 expression is decreased in breast cancer tissues and cells, and its overexpression can inhibit the invasion and metastasis of breast cancer cells by targeted regulation of HNRNPA2B1.

MiR-429 suppresses proliferation and invasion of breast cancer via inhibiting the Wnt/ß-catenin signaling pathway.

BACKGROUND: microRNAs (miRNAs) have been verified as molecular targets for regulating tumor proliferation, invasion, and metastasis in tumor progression. However, the relationship between miRNAs and cellular energy metabolism in breast cancer still needs to be clarified. This study aimed to investigate the role of miR-429 in breast cancer progression. METHODS: Bioinformatic analyses were employed to detect the relationship between miR-429 and cancer-related signaling pathways. We used a Kaplan-Meier curve to analyze survival rate in patients with high or low expression of miR-429. We used real-time quantitative PCR (RT-qPCR) to detect the expression of miR-429 in different cell lines. Sh-con, over-miR-429, miR-429 inhibitor, and sh-inhibitor control were transfected. Colony formation and EDU assay were used to detect the proliferation of transfected cells. Wound healing and transwell assays were performed to detect the mobility and invasion ability of transfected cells. Western blot assay was used to detect relative protein expression in transfected cells and different tissues. Bioinformatic analyses were conducted to detect the target proteins expression in different breast cancer databases. Dual luciferase reporter assay was used to confirm the binding site between miR-429 and fibronectin 1 (FN1). RESULTS: The results of our study indicate that MiR-429 and its target genes are associated with cancer-related signaling pathways and that higher miR-429 expression corresponds with a better prognosis. When miR-429 was overexpressed, the proliferation, invasion of MDA-MB-231 were inhibited. MiR-429 was able to suppress the Wnt/ß-catenin signaling pathway, and FN1 overexpression could rescue the influence of over-miR-429. CONCLUSIONS: The results of our study suggest that miR-429 suppresses the proliferation and invasion of breast cancer via inhibiting the Wnt/ß-catenin signaling pathway.

[miR-93-5p promotes invasion and migration of triple negative breast cancer cells by targeting FGD5].

Objective To investigate the molecular mechanism by which miR-93-5p promotes the invasion and migration of triple negative breast cancer (TNBC) cells. Methods The miR-93-5p and faciogenital dysplasia-5 (FGD5) were screened out by Gene Expression Omnibus (GEO). Bioinformatics software was used to predict the candidate target genes for miR-93-5p. The relative expression of miR-93-5p in cells was detected by real-time quantitative PCR. Western blot was used to detect the relative expression of FGD5. TranswellTM assay was performed to detect the effects of miR-93-5p on the invasion and migration of MDA-MB-231 cells. The expression of FGD5 and survival curve in breast cancer and the correlation between miR-93-5p and FGD5 were analyzed by bioinformatics database. Dual luciferase reporter gene experiment was employed to verify the targeting relationship between miR-93-5p and FGD5. Results The miR-93-5p was highly expressed in TNBC tissues and cell lines. The higher the expression of miR-93-5p was, the worse the prognosis of breast cancer patients were. Knockdown of miR-93-5p inhibited the invasion and migration ability of MDA-MB-231 cells. Bioinformatics analysis showed that there were complementary sequences between miR-93-5p and FGD5. FGD5 presented low expression in breast cancer tissues and lower FGD5 expression in breast cancer patients corresponded to poorer prognosis. The expression levels between miR-93-5p and FGD5 were negatively correlated. Transfection of miR-93-5p inhibitor plasmid up-regulated the expression of FGD5 in TNBC cells. Dual luciferase reporter gene experiments confirmed that miR-93-5p could down-regulate the luciferase activity of wild-type FGD5. Conclusion The miR-93-5p can promote the invasion and migration of TNBC cells by targeting FGD5.

[Mechanism of DERL3 Affecting the Proliferation, Invasion and Metastasis of Lung Adenocarcinoma A549 Cells].

BACKGROUND: Derlin 3 (DERL3) is downregulated in colorectal cancer (CRC) samples. Its level is closely linked to lymphatic metastasis or distant metastasis rate in CRC patients. However, its biological behavior in lung adenocarcinoma were rarely reported. The aim of this study is to investigate the ectopic expression of DERL3 in lung adenocarcinoma tissues and its effect on the invasion and metastasis of lung adenocarcinoma A549 cell line to reveal the possible mechanism of invasion and metastasis of lung adenocarcinoma. METHODS: Lung adenocarcinoma microarray gene chip data included 3 cases of lymph node metastasis and 3 cases of lung adenocarcinoma tissue without lymph node metastasis. The GEDS and Kaplan-Meier plot queries the survival curve and expression level of DERL3. Western blot was used to detect the expression of DERL3 in lung adenocarcinoma cells. The efficiency of knockdown DERL3 gene was detected by Western blot assay. Transwell detected the number of cells passing through the basement membrane of the transwell. EDU assay detected cell proliferation ability. Western blot detected the expression of epithelial-mesenchymal transition related proteins E-cadherin and Vimentin. RESULTS: The microarray gene chip results showed that compared with lung adenocarcinoma tissues without lymph node metastasis, 1,314 mRNAs in lung adenocarcinoma tissues with lymph node metastasis were up-regulated, 400 mRNAs were down (P<0.05). The expression of DERL3 increased in lung adenocarcinoma (P<0.05). The results of survival curve showed that the lung cancer patients with high expression of DERL3 with poor prognosis (P<0.05). Western blot results indicated that plasmid transfection was successful. Knockdown of DERL3 suppressed the ability of proliferation, invasion and migration in A549 cells (P<0.05). After knockdown of DERL3, the expression level of Vimentin was decreased, while E-cadherin expression increased (P<0.05). CONCLUSIONS: Knockdown of DERL3 inhibited the proliferation, invasion and metastasis of A549 cells.

Overexpression of PID1 reduces high density lipoprotein level and functionality in swine.

Phosphotyrosine interaction domain 1 (PID1), a protein with a phosphotyrosine-binding (PTB) domain, interacts with the lipoprotein receptor-related protein 1 (LRP1) to reduce the insulin sensitivity of adipocyte. Considering the role of LRP1 in lipid metabolism, we investigated the effect of PID1 on the content and biological activities of serum lipoproteins in pigs. PID1-transgenic pigs were genetated by sperm and magnetic nanoparticles-mediated method. The levels of PID1 in PID1-transgenic pig's liver were higher than that in the wild-type pig's liver. We found that serum levels of high-density lipoprotein cholesterol (HDL-C) and apolipoprotein A-I (apoA-I) were significantly reduced in PID1-transgenic pigs. On the other hand, PID1-transgenic pigs displayed increased non-HDL-C levels. Serum levels of total cholesterol and triglycerides were comparable between the PID1-transgenic and the wild-type pigs. Further, the HDL isolated from PID1-transgenic pigs showed a significant reduction in cholesterol efflux ability. In addition, serum superoxide dismutase activity of PID1-transgenic pigs was also obviously lowered compared with that of wild type pigs. In conclusion, these results suggest that PID1 might be able to adjust HDL-C levels in serum and HDL cholesterol efflux ability.

Sphingosine-1-Phosphate Improves the Biological Features of Mouse Bone Marrow-Derived EPCs Partially through PI3K/AKT/eNOS/NO Pathway.

Sphingosine-1-phosphate (S1P), a bioactive sphingolipid, is recognized as a critical regulator in physiological and pathophysiological processes of atherosclerosis (AS). However, the underlying mechanism remains unclear. As the precursor cells of endothelial cells (ECs), endothelial progenitor cells (EPCs) can prevent AS development through repairing endothelial monolayer impaired by proatherogenic factors. The present study investigated the effects of S1P on the biological features of mouse bone marrow-derived EPCs and the underlying mechanism. The results showed that S1P improved cell viability, adhesion, and nitric oxide (NO) release of EPCs in a bell-shaped manner, and migration and tube formation dose-dependently. The aforementioned beneficial effects of S1P on EPCs could be inhibited by the phosphatidylinositol 3-kinase (PI3K) inhibitor of LY294002 and nitric oxide synthase (NOS) inhibitor of N'-nitro-L-arginine-methyl ester hydrochloride (L-NAME). The inhibitor of LY294002 inhibited S1P-stimulated activation of phosphorylated protein kinase B (AKT) (p-AKT) and endothelial nitric oxide synthase (eNOS) (p-eNOS), and down-regulated the level of eNOS significantly. The results suggest that S1P improves the biological features of EPCs partially through PI3K/AKT/eNOS/NO signaling pathway.

Reverse-D-4F improves endothelial progenitor cell function and attenuates LPS-induced acute lung injury.

BACKGROUND: Patients with acute lung injury (ALI) have increased levels of pro-inflammatory mediators, which impair endothelial progenitor cell (EPC) function. Increasing the number of EPC and alleviating EPC dysfunction induced by pro-inflammatory mediators play important roles in suppressing ALI development. Because the high density lipoprotein reverse-D-4F (Rev-D4F) improves EPC function, we hypothesized that it might repair lipopolysaccharide (LPS)-induced lung damage by improving EPC numbers and function in an LPS-induced ALI mouse model. METHODS: LPS was used to induce ALI in mice, and then the mice received intraperitoneal injections of Rev-D4F. Immunohistochemical staining, flow cytometry, MTT, transwell, and western blotting were used to assess the effect of Rev-D4F on repairment of lung impairment, and improvement of EPC numbers and function, as well as the signaling pathways involved. RESULTS: Rev-D4F inhibits LPS-induced pulmonary edema and decreases plasma levels of the pro-inflammatory mediators TNF-α and ET-1 in ALI mice. Rev-D4F inhibited infiltration of red and white blood cells into the interstitial space, reduced lung injury-induced inflammation, and restored injured pulmonary capillary endothelial cells. In addition, Rev-D4F increased numbers of circulating EPC, stimulated EPC differentiation, and improved EPC function impaired by LPS. Rev-D4F also acted via a PI3-kinase-dependent mechanism to restore levels of phospho-AKT, eNOS, and phospho-eNOS suppressed by LPS. CONCLUSIONS: These findings indicate that Rev-D4F has an important vasculoprotective role in ALI by improving the EPC numbers and functions, and Rev-D4F reverses LPS-induced EPC dysfuncion partially through PI3K/AKT/eNOS signaling pathway.

C1q/TNF-related protein 9 improves the anti-contractile effects of perivascular adipose tissue via the AMPK-eNOS pathway in diet-induced obese mice.

The anti-contractile property of perivascular adipose tissue (PVAT) is abolished through an endothelium-dependent pathway in obesity. C1q/tumor necrosis factor-related protein (CTRP)9 improved endothelial function by promoting endothelium-dependent vasodilatation. The aims of this study were to investigate whether CTRP9 improves the anti-contractile effect of PVAT and protects against PVAT dysfunction in obese mice. The mice were treated with a high-fat diet with or without CTRP9 treatment. Thoracic aortas with or without PVAT (PVAT+ or PVAT-) were prepared, and concentration-dependent responses to phenylephrine were measured. Obese mice showed a significantly increased contractile response, which was suppressed by CTRP9 treatment both with and without PVAT. PVAT significantly reduced the anti-contractile effect in obese mice, which was partially restored by CTRP9 treatment. Treatment of the aortic rings (PVAT+) with inhibitors of AMP protein kinase (AMPK), Akt and endothelial nitric oxide synthase (eNOS) attenuated the beneficial effect of CTRP9 on PVAT. Similar results were observed when we pretreated the aortic rings with CTRP9 ex vivo. CTRP9 significantly enhanced the phosphorylation levels of AMPK, Akt and eNOS, and reduced superoxide production and TNF-α levels in PVAT from obese mice. Our study suggests that CTRP9 enhanced the anti-contractile effect of PVAT and improved PVAT function by activating the AMPK-eNOS pathway in obese mice.

Bone marrow mesenchymal stromal cells alleviate brain white matter injury via the enhanced proliferation of oligodendrocyte progenitor cells in focal cerebral ischemic rats.

The effects of transplanting bone marrow mesenchymal stromal cells (BMSCs) for the treatment of white matter damage are not well understood, nor are the underlying mechanisms. Recent studies showed that endogenous oligodendrocyte progenitor cells (OPCs) can be stimulated to proliferate. Therefore, we explore the effects of BMSCs transplantation on white matter damage and the proliferation of OPCs in transient focal cerebral ischemic rats. BMSCs were transplanted into a group of rats that had undergone middle cerebral artery occlusion (MCAO) 24 h after reperfusion. The ratswere examined by MRI-T2 and DTI sequencesdynamically. The proliferating cells were labeled by 5-Bromo-2'-deoxyuridine (BrdU). The effects of BMSC transplantation on neurons, axons, myelination, and proliferating OPCs were examined by Nissl staining, MBP/NF-H and BrdU/NG2 immunofluorescence staining7 days after transplantation. More Nissl-stained neuronswere found and the FA value of MRI-DTI was significantly higher in the MCAO + BMSCs group than in the MCAOgroup (both P < .01). The fold change of MBP protein was significantly higher in the MCAO + BMSCs group than in the MCAO group (P < .01); the same was true of NF-H protein. Additionally, there were more BrdU+NG2+ cells in the SVZ areas of the MCAO + BMSCs group than in the MCAO group (P < .01). BMSCs thus were shown to alleviate neuronal/axonal injury and promote the proliferation of OPCs and formation of myelin sheath, significantly alleviating white matter damage in focal cerebral ischemic rats.

[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.

Bone marrow derived mesenchymal stem cells alleviated brain injury via down-regulation of interleukin-1ß in focal cerebral ischemic rats.

Interleukin-1ß (IL-1ß) plays an important role in brain injury after focal ischemia, and bone marrow-derived mesenchymal stem cells (BMSCs) are capable of reducing the expression of IL-1ß, we investigated the effects of BMSCs transplantation on brain edema and cerebral infarction as well as the underlying mechanisms via IL-1ß. Male Sprague-Dawley rats were randomly divided into five groups: Normal + phosphate-buffered saline (PBS), middle cerebral artery occlusion (MCAO) + PBS, Normal + BMSCs, MCAO + BMSCs and MCAO + IL-1ra (an antagonist of IL-1ß). BMSCs were transplanted 24 hours after MCAO, and brain edema was evaluated by Magnetic Resonance Imaging (MRI) and brain water content method after BMSCs transplantation. The expression of NeuN and AQP4 was analyzed by immunofluorescence staining. Protein level of AQP4 and IL-1ß was detected by western blot analysis 48 hours after transplantation. The results showed that BMSCs transplantation reduced brain edema by measurement of brain water content and ADC Value of MRI, as well as the expression of AQP4 and IL-1ß. It was also found that BMSCs transplantation could alleviate the cerebral infarction volume and neuronal damage. Both the brain edema and the cerebral infarction were associated with IL-1ß expression. In conclusion, BMSCs transplantation was capable of alleviating brain edema as well as reducing cerebral infarction via down-regulation of IL-1ß expression, thus repair the injured brain in focal cerebral ischemic rats.

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.

[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.

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 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.