Low-pass genome sequencing versus chromosomal microarray analysis: implementation in prenatal diagnosis.

PURPOSE: Emerging studies suggest that low-pass genome sequencing (GS) provides additional diagnostic yield of clinically significant copy-number variants (CNVs) compared with chromosomal microarray analysis (CMA). However, a prospective back-to-back comparison evaluating accuracy, efficacy, and incremental yield of low-pass GS compared with CMA is warranted. METHODS: A total of 1023 women undergoing prenatal diagnosis were enrolled. Each sample was subjected to low-pass GS and CMA for CNV analysis in parallel. CNVs were classified according to guidelines of the American College of Medical Genetics and Genomics. RESULTS: Low-pass GS not only identified all 124 numerical disorders or pathogenic or likely pathogenic (P/LP) CNVs detected by CMA in 121 cases (11.8%, 121/1023), but also defined 17 additional and clinically relevant P/LP CNVs in 17 cases (1.7%, 17/1023). In addition, low-pass GS significantly reduced the technical repeat rate from 4.6% (47/1023) for CMA to 0.5% (5/1023) and required less DNA (50 ng) as input. CONCLUSION: In the context of prenatal diagnosis, low-pass GS identified additional and clinically significant information with enhanced resolution and increased sensitivity of detecting mosaicism as compared with the CMA platform used. This study provides strong evidence for applying low-pass GS as an alternative prenatal diagnostic test.

Characteristics and mode of inheritance of pathogenic copy number variants in prenatal diagnosis.

BACKGROUND: Microdeletions and microduplications can occur in any pregnancy independent of maternal age. The spectrum and features of pathogenic copy number variants including the size, genomic distribution, and mode of inheritance are not well studied. These characteristics have important clinical implications regarding expanding noninvasive prenatal screening for microdeletions and microduplications. OBJECTIVES: The aim was to investigate the spectrum and characteristics of pathogenic copy number variants in prenatal genetic diagnosis and to provide recommendations for expanding the scope of noninvasive prenatal screening for microdeletions and microduplications. STUDY DESIGN: This was a retrospective study of 1510 pregnant women who underwent invasive prenatal diagnostic testing by chromosomal microarray analysis. Prenatal samples were retrieved by amniocentesis or chorionic villus sampling and sent to our prenatal genetic diagnosis laboratory for chromosomal microarray analysis. The risk of carrying a fetus with pathogenic copy number variants is stratified by the patients' primary indication for invasive testing. We searched the literature for published prenatal chromosomal microarray data to generate a large cohort of 23,865 fetuses. The characteristics and spectrum of pathogenic copy number variants including the type of aberrations (gains or losses), genomic loci, sizes, and the mode of inheritance were studied. RESULTS: Overall, 375 of 23,865 fetuses (1.6%) carried pathogenic copy number variants for any indication for invasive testing, and 44 of them (11.7%) involve 2 or more pathogenic copy number variants. A total of 428 pathogenic copy number variants were detected in these fetuses, of which 280 were deletions and 148 were duplications. Three hundred sixty (84.1%) were less than 5 Mb in size and 68 (15.9%) were between 5 and 10 Mb. The incidence of carrying a pathogenic copy number variant in the high-risk group is 1 in 36 and the low-risk group is 1 in 125. Parental inheritance study results were available for 311 pathogenic copy number variants, 71 (22.8%) were maternally inherited, 36 (11.6%) were paternally inherited, and 204 (65.6%) occurred de novo. CONCLUSION: Collectively, pathogenic copy number variants are common in pregnancies. High-risk pregnancies should be offered invasive testing with chromosomal microarray analysis for the most comprehensive investigation. Detection limits on size, parental inheritance, and genomic distribution should be carefully considered before implementing copy number variant screening in expanded noninvasive prenatal screening.

EP4 emerges as a novel regulator of bile acid synthesis and its activation protects against hypercholesterolemia.

Prostaglandin E receptor subtype 4 (EP4) knockout mice develops spontaneous hypercholesterolemia but the detailed mechanisms by which EP4 affects cholesterol homeostasis remains unexplored. We sought to determine the cause of hypercholesterolemia in EP4 knockout mice, focusing on the role of EP4 in regulating the synthesis and elimination of cholesterol. Deficiency of EP4 significantly decreased total bile acid levels in the liver by 26.2% and the fecal bile acid content by 27.6% as compared to wild type littermates, indicating that the absence of EP4 decreased hepatic bile acid synthesis and their subsequent excretion in stools. EP4 deficiency negatively regulate bile acid synthesis through repression of phosphorylated extracellular signal-regulated kinase 1/2 (ERK)-mediated cholesterol 7α-hydroxylase (CYP7A1) expression and that the hypercholesterolemia in EP4 knockout mice is due to a defect in cholesterol conversion into bile acids. Deficiency of EP4 also increased de novo cholesterol synthesis and altered cholesterol fluxes in and out of the liver. Treating high fat diet-challenged mice with the pharmacological EP4 agonist, CAY10580 (200 µg/kg body weight/day i.p) for three weeks effectively prevented diet-induced hypercholesterolemia, enhanced endogenous bile acid synthesis and their fecal excretion. In summary, EP4 plays a critical role in maintaining cholesterol homeostasis by regulating the synthesis and elimination of bile acids. Activation of EP4 serves as an effective novel strategy to promote cholesterol disposal in the forms of bile acids in order to lower plasma cholesterol levels.

Histological and microRNA Signatures of Corneal Epithelium in Keratoconus.

PURPOSE: To illustrate the histopathology of keratoconic corneal epithelia and its micro-ribonucleic acid (miRNA) regulation. METHODS: Corneal epithelia were collected from 27 patients with keratoconus and 26 normal patients after surgery or by impression cytology. The miRNA profile was determined using miRNA microarray. The biological roles of miRNA target genes were delineated by gene ontology and pathway analyses. The expressions of significant miRNAs were validated using TaqMan polymerase chain reaction (PCR), whereas protein localization and expression of the miRNA target genes were examined by immunofluorescence and immunoblotting analyses. RESULTS: Histological assessment showed that corneal epithelia in patients with keratoconus were thinner with loosely packed cells compared to normal patients. Microarray analysis revealed that 12 miRNAs were significantly downregulated in keratoconic corneal epithelia. Gene ontology analysis demonstrated that the predicted miRNA target genes participated in cell junction, cell division, and motor activity, whereas pathway analysis highlighted the involvement of syndecan-mediated signaling pathway. TaqMan PCR validated the altered expression of six miRNAs in corneal epithelia from surgery (hsa-miR-151a-3p, hsa-miR-138-5p, hsa-miR-146b-5p, hsa-miR-194-5p, hsa-miR-28-5p, and hsa-miR-181a-2-3p) and four miRNAs in squamous corneal epithelial samples collected from impression cytology (hsa-miR-151a-3p, hsa-miR-195-5p, hsa-miR-185-5p, and hsa-miR-194-5p). In addition, higher S100A2 expression was found in the epithelial basal cell layer of keratoconic corneal epithelia. CONCLUSIONS: The miRNA and histological analyses in this study demonstrated structural and biological changes in keratoconic corneal epithelia, broadening the understanding of keratoconus pathology. In addition, impression cytology is useful to collect corneal epithelial tissues for gene expression analysis. [J Refract Surg. 2018;34(3):201-211.].

The CCCTC-binding factor (CTCF)-forkhead box protein M1 axis regulates tumour growth and metastasis in hepatocellular carcinoma.

CCCTC-binding factor (CTCF) is a DNA-binding protein that interacts with a large number of highly divergent target sequences throughout the genome. It is implicated in a variety of functions, including chromatin organization and transcriptional control. The functional role of CTCF in tumour pathogenesis remains elusive. We showed that CTCF is frequently upregulated in a subset of primary hepatocellular carcinomas (HCCs) as compared with non-tumoural liver. Overexpression of CTCF was associated with shorter disease-free survival of patients. Short hairpin RNA (shRNA)-mediated suppression of CTCF inhibited cell proliferation, motility and invasiveness in HCC cell lines; these effects were correlated with prominent reductions in the expression of telomerase reverse transcriptase (TERT), the shelterin complex member telomerase repeat-binding factor 1, and forkhead box protein M1 (FOXM1). In contrast, upregulation of CTCF was positively correlated with FOXM1 and TERT expression in clinical HCC biopsies. Depletion of CTCF resulted in reduced motility and invasiveness in HCC cells that could be reversed by ectopic expression of FOXM1, suggesting that FOXM1 is one of the important downstream effectors of CTCF in HCC. Reporter gene analysis suggested that depletion of CTCF is associated with reduced FOXM1 and TERT promoter activity. Chromatin immunoprecipitation (ChIP)-polymerase chain reaction (PCR) analysis further revealed occupancy of the FOXM1 promoter by CTCF in vivo. Importantly, depletion of CTCF by shRNA significantly inhibited tumour progression and metastasis in HCC mouse models. Our work uncovered a novel functional role of CTCF in HCC pathogenesis, which suggests that targeting CTCF could be further explored as a potential therapeutic strategy for HCC. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Rap1 induces cytokine production in pro-inflammatory macrophages through NFκB signaling and is highly expressed in human atherosclerotic lesions.

Repressor activator protein 1 (Rap1) is essential for maintaining telomere length and structural integrity, but it also exerts other non-telomeric functions. The present study tested the hypothesis that Rap1 is released into the cytoplasm and induces production of pro-inflammatory cytokines via nuclear factor kappa B (NFκB) signaling in macrophages, a cell type involved in the development and progression of atherosclerotic lesions. Western blotting analysis confirmed that Rap1 was present in the cytoplasm of differentiated human monocytic leukemia cells (THP-1, a macrophage-like cell line). Co-immunoprecipitation assay revealed a direct interaction between Rap1 and I kappa B kinase (IKK). Knockdown of Rap1 suppressed lipopolysaccharide-mediated activation of NFκB, and phosphorylation of inhibitor of kappa B α (IκBα) and p65 in THP-1 macrophages. The reduction of NFκB activity was paralleled by a decreased production of NFκB-dependent pro-inflammatory cytokines and an increased expression of IκBα (native NFκB inhibitor) in various macrophage models with pro-inflammatory phenotype, including THP-1, mouse peritoneal macrophages and bone marrow-derived M1 macrophages. These changes were observed selectively in pro-inflammatory macrophages but not in bone marrow-derived M2 macrophages (with an anti-inflammatory phenotype), mouse lung endothelial cells, human umbilical vein endothelial cells or human aortic smooth muscle cells. Immunostaining revealed that Rap1 was localized mainly in macrophage-rich areas in human atherosclerotic plaques and that the presence of Rap1 was positively correlated with the advancement of the disease process. In pro-inflammatory macrophages, Rap1 promotes cytokine production via NFκB activation favoring a pro-inflammatory environment which may contribute to the development and progression of atherosclerosis.

Signature microRNAs in human cornea limbal epithelium.

This study was aimed to identify the signature microRNAs, which regulate the biological processes of corneal epithelial progenitor cell (CEPC) homeostasis and regulation through characterizing the differential expression profile of microRNAs in human limbal epithelium containing adult CEPC versus central corneal epithelium without CEPC. MicroRNA microarray had identified 37 microRNAs enriched in human corneal epithelium. Among them, nine were significantly upregulated in limbal epithelium and one in central corneal epithelium after validation by TaqMan® real-time polymerase chain reaction. In addition to our previous finding of miR-143 and 145, the expression of miR-10b, 126, and 155 was localized in limbal epithelium (LE) (predominantly basal layers) by using locked nucleic acid-based in situ hybridization. Potential target genes were predicted by TargetScan Human v6.0 and compared to the reported human cornea epithelial gene profile GSE5543. Analyzed by web-based Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and DAVID Functional Annotation Bioinformatics Resources v6.7, the downregulated genes were involved in pathways of immune response and cellular protection, apoptosis, and cell movement whereas upregulated genes with cell survival, cell-matrix interaction, and cell-cell adhesion. We found a constant occurrence of miR-143, 145, and 155 in all KEGG pathways regulating limbal epithelial events. By Ingenuity Systems (IPA®) analysis, these microRNAs could cooperatively regulate cell growth and apoptosis via tumor necrosis factor activation and MYC repression. Our findings thus suggest a unique microRNA signature existing in human limbal epithelium and participating in CEPC homeostasis.

Adrenomedullin and diabetes.

Adrenomedullin (ADM) is a peptide hormone widely expressed in different tissues, especially in the vasculature. Apart from its vasodilatatory and hypotensive effect, it plays multiple roles in the regulation of hormonal secretion, glucose metabolism and inflammatory response. ADM regulates insulin balance and may participate in the development of diabetes. The plasma level of ADM is increased in people with diabetes, while in healthy individuals the plasma ADM concentration remains low. Plasma ADM levels are further increased in patients with diabetic complications. In type 1 diabetes, plasma ADM level is correlated with renal failure and retinopathy, while in type 2 diabetes its level is linked with a wider range of complications. The elevation of ADM level in diabetes may be due to hyperinsulinemia, oxidative stress and endothelial injury. At the same time, a rise in plasma ADM level can trigger the onset of diabetes. Strategies to reduce ADM level should be explored so as to reduce diabetic complications.

Utilization of glucose, blood pressure, and lipid lowering medications among people with type II diabetes in the United States, 1999-2010.

PURPOSE: Changes in relation to drug treatment to various control targets for diabetes were studied using the National Health and Nutrition Examination Survey, 1999-2010. METHODS: Data on 3094 participants aged 20 years or older with diagnosed type II diabetes were analyzed. Use of medications for lowering glucose, blood pressure, and lipids in the past month was assessed by questionnaire. Data from two survey cycles were combined together to produce estimates for each 4-year period. RESULTS: Usage of metformin increased from 34.8% to 53.8% and was the most prevalent medications during this period (P < .001), and half of subjects taking metformin could achieve glycated hemoglobin less than 7.0% in 2007-2010. Dipeptidyl peptidase-4 inhibitors were used by 7.4% of participants in 2007-2010. Usage of angiotensin receptor blockers and beta-blockers increased significantly from 7.4% to 21.4% and from 15.3% to 31.8%, respectively from 1999 to 2010 (P ≤ .001). A total of 64.7% of participants could attain blood pressure control by 2007-2010. Usage of statins doubled in 1999-2010 and 52.2% of subjects took statins by 2007-2010 (P < .001). CONCLUSIONS: Metformin is the first-line drug for diabetes while dipeptidyl peptidase-4 inhibitors started to be used since 2007. Blood pressure control improved in 1999-2010 partly due to increased drug prescriptions. Although statins were widely used about half of the participants did not take them.

Plasma level of adrenomedullin is influenced by a single nucleotide polymorphism in the adiponectin gene.

OBJECTIVE: Adrenomedullin (ADM) and adiponectin are both involved in inflammation and cardiovascular diseases. The plasma levels of these peptides are influenced by single nucleotide polymorphisms (SNPs) in the ADM and ADIPOQ genes respectively. There is some evidence that ADM may regulate adiponectin gene expression, but whether adiponectin can regulate ADM expression is unclear, and was therefore investigated. METHODS: Plasma ADM level was measured in 476 subjects in the Hong Kong Cardiovascular Risk Factor Prevalence Study-2 (CRISPS2). We genotyped them for 2 ADIPOQ SNPs that are known to be associated with plasma adiponectin level. RESULTS: The minor allele frequencies of ADIPOQ SNPs rs182052 and rs12495941 were 40.6% and 42.2% respectively. Plasma ADM level was significantly associated with rs182052 after adjusting for age and sex (ß=0.104, P=0.023) but not with rs12495941 (ß=0.071, P=0.120). In multivariate analysis, plasma ADM level increased with the number of minor alleles of rs182052 (P=0.013). Compared to subjects with GG genotype, subjects with AA genotype had 17.7% higher plasma ADM level (95% CI: 3.6%-33.7%). Subgroup analysis revealed that the association was significant in diabetic patients (ß=0.344, P=0.001) but not in non-diabetic subjects. CONCLUSION: Plasma ADM level is related to SNP rs182052 in the ADIPOQ gene. Our findings provide new evidence of the interplay between these two important peptides in cardiovascular disease and diabetes. Knowing the genotype may help to refine the interpretation of these biomarkers.

MicroRNA-182 plays an onco-miRNA role in cervical cancer.

OBJECTIVES: The purposes of this study were to identify aberrantly expressed miRNAs and investigate their pathogenic roles in cervical cancer. METHODS: miRNA expression was assessed in cervical cancer cell lines, micro-dissected normal cervical epithelium cells and primary cervical carcinoma by TaqMan RT-PCR. Spatial expression of miR-182 in cervical carcinoma and normal cervix was explored by in situ hybridization. HeLa xenograft mice model was used for evaluation of the effect on tumor growth of miR-182 inhibitor. Western blot, flow cytometry and gene expression analysis were used for identification of the functional role of miR-182 in HeLa cells. RESULTS: Two up-regulated (miR-182 and -183) and nine down-regulated (miR-211, 145, 223, 150, 142-5p, 328, 195, 199b, 142-3p) microRNAs were consistently identified in cervical cancer cell lines. Further investigation confirmed the most up-regulated miRNA (miR-182) was significantly elevated in primary cervical carcinoma and discovered a significant correlation between the increased expression of miR-182 and advanced stages of cervical cancer. In HeLa xenograft mouse model, we demonstrated that inhibition of the miR-182 could exert the effect of tumor growth regression. Western blot, flow cytometry and pathway analysis for the HeLa cells with miR-182 over/down-expression in vitro showed that miR-182 was involved in apoptosis and cell cycle pathways, it also associated with the regulation of FOXO1. CONCLUSIONS: Our findings indicated that miR-182 plays an onco-miRNA role in cervical cancer and its alteration is associated with cervical cancer pathogenesis by disrupting cell proliferation.

A single nucleotide polymorphism of interleukin-6 gene is related to plasma adrenomedullin levels.

OBJECTIVE: Elevated plasma adrenomedullin (ADM) levels are associated with cardiovascular diseases. Single nucleotide polymorphisms (SNPs) in the gene encoding ADM (ADM) are associated with plasma ADM levels. The presence of a nuclear factor for interleukin-6 (IL-6) expression binding site in the promoter region of the ADM gene suggests a possible relationship between the expression of the ADM and IL-6. Therefore, we investigated whether plasma ADM levels are related to SNPs in the gene encoding IL-6 (IL6). METHODS: Plasma ADM levels were measured in 476 subjects in the Hong Kong Cardiovascular Risk Factor Prevalence Study-2 (CRISPS2). The subjects were genotyped for three tagging SNPs in the IL6 gene. RESULTS: The minor allele frequencies of the IL6 SNPs rs17147230, rs1800796 and rs2069837 were 41·8%, 20·0% and 15·4%, respectively. The tagging SNP, rs17147230, was associated with plasma ADM levels after adjusting for age and sex (ß=-0·096, P = 0·034). The association was significant in women (ß=-0·115, P = 0·021) but not in men. Among all subjects, plasma ADM levels decreased with an increasing number of minor alleles of rs17147230 in multivariate analysis (P = 0·034). Compared to subjects with the AA genotype, subjects with the TT genotype had plasma ADM levels 12·8% lower (95% CI: 0·6-23·5%, P = 0·041). Haplotype analysis demonstrated a significant association of the haplotype ACA with plasma ADM levels in women (P < 0·05). CONCLUSION: Plasma ADM levels are related to the SNP rs17147230 in IL6 gene. The effect of the polymorphism on inflammation and cardiovascular disease remains to be determined.

The detection of mosaicism by prenatal BoBs™.

OBJECTIVE: The objective of the study was to evaluate the ability of a new prenatal diagnostic platform - prenatal BACs-on-Beads™ (BoBs™) in detecting mosaicism by comparison to quantitative fluorescence-polymerase chain reaction (QF-PCR). METHODS: A validation study of prenatal BoBs™ was firstly performed using 18 artificially constructing mosaic samples involving various aneuploidies and microdeletion conditions. Additionally, we compared the accuracy between prenatal BoBs™ and QF-PCR for 18 archived clinical mosaic cases and nine chromosomally abnormal cell lines with reference to conventional karyotype results. RESULTS: In the validation study, BoBs™ allowed the detection of mosaicism at a level of 20-40%. Among the clinical mosaic cases, 14/18 cases were within the detection of BoBs™, 8/14 (57.1%) could be identified by BoBs™ and 6/9 (66.7%) by QF-PCR, but 6/14 (42.9%) were missed by both tests. Three cases (16.7%) were detected by prenatal BoBs™ but missed by QF-PCR, whereas QF-PCR detected one case that was missed by BoBs™. The overall sensitivity of BoBs™ in detecting mosaicism is 44.4% (8/18), which is slightly higher than that of QF-PCR (33.3%; 6/18). CONCLUSION: Prenatal BoBs™ has a sensitivity of 57.1% in the detection clinical mosaic cases. According to the validation test, mosaicism of 20% or greater is detectable by the BoBs™ assay.

Nicotine alters MicroRNA expression and hinders human adult stem cell regenerative potential.

Adult stem cells are critical for the healing process in regenerative medicine. However, cigarette smoking inhibits stem cell recruitment to tissues and delays the wound-healing process. This study investigated the effect of nicotine, a major constituent in the cigarette smoke, on the regenerative potentials of human mesenchymal stem cells (MSC) and periodontal ligament-derived stem cells (PDLSC). The cell proliferation of 1.0 µM nicotine-treated MSC and PDLSC was significantly reduced when compared to the untreated control. Moreover, nicotine also retarded the locomotion of these adult stem cells. Furthermore, their osteogenic differentiation capabilities were reduced in the presence of nicotine as evidenced by gene expression (RUNX2, ALPL, BGLAP, COL1A1, and COL1A2), calcium deposition, and alkaline phosphatase activity analyses. In addition, the microRNA (miRNA) profile of nicotine-treated PDLSC was altered; suggesting miRNAs might play an important role in the nicotine effects on stem cells. This study provided the possible mechanistic explanations on stem cell-associated healing delay in cigarette smoking.

Is human cytomegalovirus infection associated with hypertension? The United States National Health and Nutrition Examination Survey 1999-2002.

PURPOSE: Recent studies have implicated the human cytomegalovirus (HCMV) as a possible pathogen for causing hypertension. We aimed to study the association between HCMV infection and hypertension in the United States National Health and Nutrition Examination Survey (NHANES). METHODS: We analyzed data on 2979 men and 3324 women in the NHANES 1999-2002. We included participants aged 16-49 years who had valid data on HCMV infection and hypertension. RESULTS: Of the participants, 54.7% had serologic evidence of HCMV infection and 17.5% had hypertension. There were ethnic differences in the prevalence of HCMV infection (P<0.001) and hypertension (P<0.001). The prevalence of both increased with age (P<0.001). Before adjustment, HCMV seropositivity was significantly associated with hypertension in women (OR=1.63, 95% CI=1.25-2.13, P=0.001) but not in men. After adjustment for race/ethnicity, the association between HCMV seropositivity and hypertension in women remained significant (OR=1.55, 95% CI=1.20-2.02, P=0.002). Further adjustment for body mass index, diabetes status and hypercholesterolemia attenuated the association (OR=1.44, 95% CI=1.10-1.90, P=0.010). However, after adjusting for age, the association was no longer significant (OR=1.24, 95% CI=0.91-1.67, P=0.162). CONCLUSIONS: In this nationally representative population-based survey, HCMV seropositivity is associated with hypertension in women in the NHANES population. This association is largely explained by the association of hypertension with age and the increase in past exposure to HCMV with age.

Adrenomedullin and cardiovascular diseases.

The cardiovascular system is regulated by the autonomic nervous system, the renin-angiotensin-aldosterone system, nitric oxide (NO) and other factors including neuropeptides. Research in neurohumoral factors has led to the development of many cardiovascular drugs. Adrenomedullin (ADM), initially isolated from the adrenal gland, has diverse physiological and pathophysiological functions in the cardiovascular system. It is produced in many organs and tissues including the vasculature. ADM has numerous actions, including vasodilation, natriuresis, antiapoptosis and stimulation of NO production. It might play a protective role in various cardiovascular pathologies, and its plasma level is elevated in patients with hypertension and heart failure. Administration of ADM is a possible therapeutic approach for treating cardiovascular diseases. A number of studies have investigated the infusion of ADM in humans, which seems to be benficial in heart failure and myocardial infarction. Instead of ADM infusion, augmentation of its endogenous level is another possible strategy. Gene therapy is feasible in animal models, but its application in humans is limited. At present, the most promising clinical application of ADM is the use of the plasma level of mid-regional proadrenomedullin as a biomarker in cardiovascular diseases. It is a good marker of prognosis and survival in patients with coronary aretery disease or heart failure.

MicroRNA-145 regulates human corneal epithelial differentiation.

BACKGROUND: Epigenetic factors, such as microRNAs, are important regulators in the self-renewal and differentiation of stem cells and progenies. Here we investigated the microRNAs expressed in human limbal-peripheral corneal (LPC) epithelia containing corneal epithelial progenitor cells (CEPCs) and early transit amplifying cells, and their role in corneal epithelium. METHODOLOGY/PRINCIPAL FINDINGS: Human LPC epithelia was extracted for small RNAs or dissociated for CEPC culture. By Agilent Human microRNA Microarray V2 platform and GeneSpring GX11.0 analysis, we found differential expression of 18 microRNAs against central corneal (CC) epithelia, which were devoid of CEPCs. Among them, miR-184 was up-regulated in CC epithelia, similar to reported finding. Cluster miR-143/145 was expressed strongly in LPC but weakly in CC epithelia (P = 0.0004, Mann-Whitney U-test). This was validated by quantitative polymerase chain reaction (qPCR). Locked nucleic acid-based in situ hybridization on corneal rim cryosections showed miR-143/145 presence localized to the parabasal cells of limbal epithelium but negligible in basal and superficial epithelia. With holoclone forming ability, CEPCs transfected with lentiviral plasmid containing mature miR-145 sequence gave rise to defective epithelium in organotypic culture and had increased cytokeratin-3/12 and connexin-43 expressions and decreased ABCG2 and p63 compared with cells transfected with scrambled sequences. Global gene expression was analyzed using Agilent Whole Human Genome Oligo Microarray and GeneSpring GX11.0. With a 5-fold difference compared to cells with scrambled sequences, miR-145 up-regulated 324 genes (containing genes for immune response) and down-regulated 277 genes (containing genes for epithelial development and stem cell maintenance). As validated by qPCR and luciferase reporter assay, our results showed miR-145 suppressed integrin ß8 (ITGB8) expression in both human corneal epithelial cells and primary CEPCs. CONCLUSION/SIGNIFICANCE: We found expression of miR-143/145 cluster in human corneal epithelium. Our results also showed that miR-145 regulated the corneal epithelium formation and maintenance of epithelial integrity, via ITGB8 targeting.