MicroRNA-146a Induces Lineage-Negative Bone Marrow Cell Apoptosis and Senescence by Targeting Polo-Like Kinase 2 Expression.

OBJECTIVE: Lineage-negative bone marrow cells (lin- BMCs) are enriched in endothelial progenitor cells and mediate vascular repair. Aging-associated senescence and apoptosis result in reduced number and functionality of lin- BMCs, impairing their prorepair capacity. The molecular mechanisms underlying lin- BMC senescence and apoptosis are poorly understood. MicroRNAs (miRNAs) regulate many important biological processes. The identification of miRNA-mRNA networks that modulate the health and functionality of lin- BMCs is a critical step in understanding the process of vascular repair. The aim of this study was to characterize the role of the miR-146a-Polo-like kinase 2 (Plk2) network in regulating lin- BMC senescence, apoptosis, and their angiogenic capability. APPROACH AND RESULTS: Transcriptome analysis in lin- BMCs isolated from young and aged wild-type and ApoE-/- (apolipoprotein E) mice showed a significant age-associated increase in miR-146a expression. In silico analysis, expression study and Luciferase reporter assay established Plk2 as a direct target of miR-146a. miR-146a overexpression in young lin- BMCs inhibited Plk2 expression, resulting in increased senescence and apoptosis, via p16Ink4a/p19Arf and p53, respectively, as well as impaired angiogenic capacity in vitro and in vivo. Conversely, suppression of miR-146a in aged lin- BMCs increased Plk2 expression and rejuvenated lin- BMCs, resulting in decreased senescence and apoptosis, leading to improved angiogenesis. CONCLUSIONS: (1) miR-146a regulates lin- BMC senescence and apoptosis by suppressing Plk2 expression that, in turn, activates p16Ink4a/p19Arf and p53 and (2) modulation of miR-146a or its target Plk2 may represent a potential therapeutic intervention to improve lin- BMC-mediated angiogenesis and vascular repair.

MicroRNA-10A* and MicroRNA-21 modulate endothelial progenitor cell senescence via suppressing high-mobility group A2.

RATIONALE: Endothelial progenitor cells (EPCs) contribute to the regeneration of endothelium. Aging-associated senescence results in reduced number and function of EPCs, potentially contributing to increased cardiac risk, reduced angiogenic capacity, and impaired cardiac repair effectiveness. The mechanisms underlying EPC senescence are unknown. Increasing evidence supports the role of microRNAs in regulating cellular senescence. OBJECTIVE: We aimed to determine whether microRNAs regulated EPC senescence and, if so, what the underlying mechanisms are. METHODS AND RESULTS: To map the microRNA/gene expression signatures of EPC senescence, we performed microRNA profiling and microarray analysis in lineage-negative bone marrow cells from young and aged wild-type and apolipoprotein E-deficient mice. We identified 2 microRNAs, microRNA-10A* (miR-10A*), and miR-21, and their common target gene Hmga2 as critical regulators for EPC senescence. Overexpression of miR-10A* and miR-21 in young EPCs suppressed Hmga2 expression, caused EPC senescence, as evidenced by senescence-associated ß-galactosidase upregulation, decreased self-renewal potential, increased p16(Ink4a)/p19(Arf) expression, and resulted in impaired EPC angiogenesis in vitro and in vivo, resembling EPCs derived from aged mice. In contrast, suppression of miR-10A* and miR-21 in aged EPCs increased Hmga2 expression, rejuvenated EPCs, resulting in decreased senescence-associated ß-galactosidase expression, increased self-renewal potential, decreased p16(Ink4a)/p19(Arf) expression, and improved EPC angiogenesis in vitro and in vivo. Importantly, these phenotypic changes were rescued by miRNA-resistant Hmga2 cDNA overexpression. CONCLUSIONS: miR-10A* and miR-21 regulate EPC senescence via suppressing Hmga2 expression and modulation of microRNAs may represent a potential therapeutic intervention in improving EPC-mediated angiogenesis and vascular repair.

[Analysis of the mutations of BRCA1 in 9 familiar breast cancer patients].

OBJECTIVE: To analyze the mutations of BRCA1 in 9 Chinese familiar breast cancer patients. METHODS: Peripheral blood samples were obtained from 9 patients enrolled from 9 breast cancer families, one normal control, 32 sporadic breast cancer patients and 33 normal donors. DNA extracted from lymphocytes was amplified by polymerase chain reaction (PCR). The 22 exons and partial introns of BRCA1 were screened by PCR-denaturing high performance liquid chromatography (PCR-DHPLC) and confirmed by direct sequencing. RESULTS: Among these 9 familiar breast cancer patients, a deleterious mutation was detected in one case in exon 11 (3870delTGTC) which was a 4-base deletion and caused a frameshift in turn. One novel and unique amino acid substitution (E867R) was detected in one case. Eight patients were detected to have a known variation in intron 18 (IVS18 + 65G-->A), and the ratio of this variation detected was 88.9% (8/9). The ratio of this variation was 37.5% (12/32) in sporadic breast cancer patients or 33.3% (11/33) in normal control. This variation was found to be accompanied all the time with a known missense variation in exon 11 (P871L) and a polymorphism in intron 9 (IVS8-57delT). Those three variants were also detected in homozygous in one case, which implies the linkage of the 3 sites. The linkage had not been reported. Two patients had been found with a known polymorphism in exon 13 (S1436S). Another known polymorphism was found in one case (L771L). In addition, intronic variants (IVS2 + 48C-->T, IVS2 + 133C-->T, IVS12 + 112C-->A) were detected. CONCLUSION: The mutations of BRCA1 in Chinese familiar breast cancer patients are different from the hot spots reported in Caucasian and Jewish. It is important that further study be conducted to seek for specific mutations of this gene or other possible relevant genes in Chinese familiar breast cancer patients.

Human umbilical cord blood endothelial progenitor cells decrease vein graft neointimal hyperplasia in SCID mice.

AIMS: Vein graft endothelial damage is a key step in the development of neointimal hyperplasia, leading to vein graft failure. We sought to determine whether exogenous endothelial progenitor cells could promote vein graft re-endothelialization, and thereby ameliorate neointimal hyperplasia. METHODS AND RESULTS: Carotid artery interposition grafting was performed with syngeneic inferior vena cavae in mice with severe combined immunodeficiency (SCID). Lineage-negative human umbilical cord blood (hUCB) cells (or medium alone) were injected into vein-grafted mice intra-operatively and 2 weeks post-operatively. In vein grafts from hUCB cell-injected mice, we found human HLA-expressing endothelial cells, as well as increased levels of VEGF and FGF-2. Furthermore, hUCB cells secreted VEGF and FGF-2 in vitro. The markedly enhanced endothelial regeneration, likely resulting from both direct engraftment and paracrine actions of hUCB cells, inhibited inflammatory response, diminished intimal cell proliferation, and reduced neointimal hyperplasia in the vein grafts. CONCLUSIONS: hUCB cells may accelerate vein graft re-endothelialization via both direct differentiation into endothelial cells and release of paracrine factors to enhance endothelial regeneration and reduce inflammation. These data highlight a potential therapeutic role for cellular therapy in vessel injury.