Minglin Ou

Hematopoietic and mesenchymal stem cell transplantation for severe and refractory systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease that is characterized by multi-organ involvement leading to significant morbidity and mortality in predominantly young women. The underlying pathogenesis involves the emergence of autoreactive T and B lymphocytes, production of autoantibodies, formation and deposition of immune complexes in various tissues leading to inflammation and organ damage. Recently, growing evidence suggests that the functions of hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs) are disrupted in SLE pathology. And HSC or MSC transplantation (HSCT/MSCT) can offer an effective and safe therapy for the severe SLE patients, resulting in disease clinical remission and improvement of organ dysfunction. In this article, we provide a brief overview of current research of autologous or allogeneic HSCT/MSCT in SLE and describe our current understanding of the mechanisms by which it plays a part in treating SLE, for better understanding of the pathogenesis, diagnosis and treatment for SLE.

Comparison of immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) assessment for Her-2 status in breast cancer.

BackgroundThe concordance rate between IHC and FISH according to clinical performance is still controversial. We report a prospective study to reflect the concordance between IHC and FISH in Guilin city, Peoples Republic of China.MethodsFifty cases of invasive ductal carcinoma of breast tested by IHC and scored as 0, 1+, 2+ and 3+ by pathologists were further analyzed by FISH using a commercially available double-color probe, and the FISH findings were compared with IHC test results.ResultsA total concordance of 82.0% was observed with a Kappa coefficient of 0.640 (P < 0.001). A high discordance was observed in 30.0% of the patients with IHC 2+, 7.1% in IHC 3+, 19.2% overall in IHC 0 and 1+.ConclusionThe IHC can be used firstly to screen the HER-2 status, and FISH can be used as a supplementary role to IHC and 2+ and some negative cases. And only those cases with Her-2 status of IHC 3+ or FISH positive should be treated with Herceptin.

Circular RNA and gene expression profiles in gastric cancer based on microarray chip technology

The aim of the present study was to screen gastric cancer (GC) tissue and adjacent tissue for differences in mRNA and circular (circRNA) expression, to analyze the differences in circRNA and mRNA expression, and to investigate the circRNA expression in gastric carcinoma and its mechanism. circRNA and mRNA differential expression profiles generated using Agilent microarray technology were analyzed in the GC tissues and adjacent tissues. qRT-PCR was used to verify the differential expression of circRNAs and mRNAs according to the interactions between circRNAs and miRNAs as well as the possible existence of miRNA and mRNA interactions. We found that: i) the circRNA expression profile revealed 1,285 significant differences in circRNA expression, with circRNA expression downregulated in 594 samples and upregulated in 691 samples via interactions with miRNAs. The qRT-PCR validation experiments showed that hsa_circRNA_400071, hsa_circRNA_000543 and hsa_circRNA_001959 expression was consistent with the microarray analysis results. ii) 29,112 genes were found in the GC tissues and adjacent tissues, including 5,460 differentially expressed genes. Among them, 2,390 differentially expressed genes were upregulated and 3,070 genes were downregulated. Gene Ontology (GO) analysis of the differentially expressed genes revealed these genes involved in biological process classification, cellular component classification and molecular function classification. Pathway analysis of the differentially expressed genes identified 83 significantly enriched genes, including 28 upregulated genes and 55 downregulated genes. iii) 69 differentially expressed circRNAs were found that might adsorb specific miRNAs to regulate the expression of their target gene mRNAs. The conclusions are: i) differentially expressed circRNAs had corresponding miRNA binding sites. These circRNAs regulated the expression of target genes through interactions with miRNAs and might become new molecular biomarkers for GC in the future. ii) Differentially expressed genes may be involved in the occurrence of GC via a variety of mechanisms. iii) CD44, CXXC5, MYH9, MALAT1 and other genes may have important implications for the occurrence and development of GC through the regulation, interaction, and mutual influence of circRNA-miRNA-mRNA via different mechanisms.

Identification of potential microRNA-target pairs associated with osteopetrosis by deep sequencing, iTRAQ proteomics and bioinformatics.

MicroRNAs aberrantly express in many human diseases including some metabolic bone disorders. They have been found to be associated with osteoclast differentiation and function, which makes them attractive candidates for the therapy of bone. However, the potential clinical application of microRNAs in therapeutics rests heavily upon our in-depth understanding of microRNAs and their targets. To identify potential microRNA–target pairs associated with osteopetrosis, we performed a system approach including deep sequencing, iTRAQ quantitative proteomics, and bioinformatics in the peripheral blood mononuclear cells (PBMCs) taken from patients with osteopetrosis and health donors. Notably, 123 differently expressed microRNAs, 173 differently expressed proteins, and 117 computationally predicted microRNA–target pairs with reciprocally expressed level in PBMCs were found in the two sample groups. Functional annotation identified that the microRNA–target pairs were involved in cell growth, differentiation, cellular signaling network, and the network highlighted the microRNA–target pair of has-miR-320a and ADP ribosylation factor 1 (Arf1) potentially associated with CLCN7 mutations in osteopetrosis. The pair of has-miR-320a and Arf1 was further verified by real-time PCR, western blot, and the interaction between has-miR-320a and its targeted sequence on the Arf1 mRNAs was confirmed by luciferase assay. Collectively, the present study established a new system approach for the investigation of microRNAs, and the microRNA–target pairs, particular has-miR-320a and Arf1, may have important roles in osteopetrosis.

Gain of the human telomerase RNA gene TERC at 3q26 is strongly associated with cervical intraepithelial neoplasia and carcinoma.

This study investigated the gain of the human telomerase RNA gene TERC at 3q26 in patients with uterine cervix disease in the southern part of China and assessed the relationship between TERC gain and cervical pathological findings. One hundred ten cervical specimens, which were collected from patients with various kinds of uterine cervix disease that was subsequently diagnosed as chronic cervicitis and with examination results negative for intraepithelial lesion or malignancy (NILM, n = 23), mild dysplasia (cervical intraepithelial neoplasia type 1 [CIN1], n = 37), moderate dysplasia (CIN2, n = 12), severe dysplasia (CIN3, n = 10), and squamous cell carcinoma (SCA, n = 28) confirmed by histologic diagnosis, were analyzed for the proportion of abnormal cells with TERC gain using a commercially available 2-color fluorescence in situ hybridization (FISH) probe. The cases with a higher proportion of abnormal cells than the threshold evaluated by NILM were recorded as positive TERC gain. The &khgr;2 and Kruskal-Wallis tests were used to assess the associations between FISH findings and diagnoses. The incidence of positive TERC gain in the cases diagnosed as NILM or CIN1 was significantly lower than that in cases diagnosed as CIN2, CIN3, or SCA (P < 0.01). In addition, a significantly higher proportion of abnormal cells with TERC gain was found as a pathological change from CIN1 to SCA (P < 0.01). We conclude that the TERC gain seems to be an important associated genetic event in CIN and carcinoma; FISH is a potential tool for the diagnoses of uterine cervix disease.

Differential proteomic analysis of renal tissue in lupus nephritis using iTRAQ reagent technology

In clinical practice, it is difficult to monitor the repeating relapse in patients suffering from systemic lupus erythematosus (SLE), who usually associated with some potential complications, for example, lupus nephritis (LN), repetition renal biopsy is necessary to determine LN flares. To identify and quantify the total proteins in renal tissue of LN patients, isobaric tags for relative and absolute quantification (iTRAQ) technology was performed. Eight-plex iTRAQ coupled with multiple chromatographic fractionation and tandem mass spectrometry were used to analyze total proteins in renal tissue of LN patients and healthy controls. Proteins were identified by mascot, which expressed differentially were noted. A total of 490 distinct proteins were identified, 113 proteins were up-regulation or down-regulation at one fold or more alteration in levels. Among of them, there was significant deviation of four proteins between our present iTRAQ study, which are up-regulated heterogeneous nuclear ribonucleoprotein (hnRNP-), Annexins and down-regulated Argininosuccinate synthetase (ASS), aldolase. iTRAQ-based quantitative proteomic technology is efficiently applicable for identification and relative quantitation of proteome of renal tissue. Differentially expressed proteome profiles of LN patients are determined. And further investigation is necessary using large cohorts of patient samples with long-term clinical follow-up data, to assess the usefulness of the pathogenesis and novel biomarker candidates of LN, which may develop a new way for diagnosis of LN.

Genome-wide analysis of 5-hmC in the peripheral blood of systemic lupus erythematosus patients using an hMeDIP-chip

Systemic lupus erythematosus (SLE) is a chronic, potentially fatal systemic autoimmune disease characterized by the production of autoantibodies against a wide range of self-antigens. To investigate the role of the 5-hmC DNA modification with regard to the onset of SLE, we compared the levels 5-hmC between SLE patients and normal controls. Whole blood was obtained from patients, and genomic DNA was extracted. Using the hMeDIP-chip analysis and validation by quantitative RT-PCR (RT-qPCR), we identified the differentially hydroxymethylated regions that are associated with SLE. There were 1,701 genes with significantly different 5-hmC levels at the promoter region in the SLE patients compared with the normal controls. The CpG islands of 3,826 genes showed significantly different 5-hmC levels in the SLE patients compared with the normal controls. Out of the differentially hydroxymethylated genes, three were selected for validation, including TREX1, CDKN1A and CDKN1B. The hydroxymethylation levels of the three genes were confirmed by RT-qPCR. The results suggested that there were significant alterations of 5-hmC in SLE patients. Thus, these differentially hydroxymethylated genes may contribute to the pathogenesis of SLE. These findings show the significance of 5-hmC as a potential biomarker or promising target for epigenetic-based SLE therapies.

Microarray Technology for Analysis of MicroRNA Expression in Renal Biopsies of Lupus Nephritis Patients

Systemic lupus erythematosus (SLE) is a complex autoimmune disease, which predominantly occurs in females and is characterized by autoantibody production against a host of nuclear self-antigens and deposition of proinflammatory immune complexes in the organs including kidney glomeruli. MicroRNAs are small noncoding intracellular RNAs that modulate gene expression at the posttranslational level. Microarray technology is in widespread use for analysis of microRNA (miRNA) gene expression because of its flexibility and accurate high throughput. RNA microarray technology is based on nucleic acid hybridization between a mixture of labeled RNA targets and their corresponding complementary probes. This article offers a technological overview of microarray technology for analysis of microRNA gene expression in kidney biopsies from SLE patients.

Rapid gene identification in a Chinese osteopetrosis family by whole exome sequencing.

Osteopetrosis is a rare genetically heterogeneous disorder of bone metabolism characterized by increased skeleton density. In the past, standard methods for genetic diagnosis of osteopetrosis have primarily been performed by candidate gene screening and positional cloning. However, these methods are time and labor consumptive; and the genetic basis of approximately 30% of the cases is yet to be elucidated. Here, we employed whole exome sequencing of two affected individuals from an osteopetrosis family to identify a candidate mutation in CLCN7 (Y99C). It was identified from a total of 1757 and 1728 genetic variations found in either patient, which were then distilled using filtering strategies and confirmed using Sanger sequencing. We identified this mutation in six family members, while not in population matched controls. This mutation was previously found in osteopetrosis patients by other researchers. Our evolutionary analysis also indicated that it is under extremely high selective pressure, and is likely to be critical for the correct function of ClC-7, and thus is likely to be the responsible cause of disease. Collectively, our data further indicated that mutation (Y99C) may be a cause of osteopetrosis, and highlights the use of whole exome sequencing as a valuable approach to identifying disease mutations in a cost and time efficient manner.

CCDC40 mutation as a cause of primary ciliary dyskinesia: a case report and review of literature.

Primary ciliary dyskinesia (PCD) is a rare, genetically heterogeneous disorder. Genetic defects affecting motility of cilia and flagella cause chronic destructive airway disease, situs inversus and, frequently, male infertility in PCD. To date, although several genes have been implicated in PCD, the genetic bases of most cases of PCD remain elusive.