Biomedical subjects

Zhenwu Lin

Publications and source records attributed to Zhenwu Lin.

6 recordsLinked to original sources

PubMed2006

High-throughput DNA methylation profiling using universal bead arrays.

We have developed a high-throughput method for analyzing the methylation status of hundreds of preselected genes simultaneously and have applied it to the discovery of methylation signatures that distinguish normal from cancer tissue samples. Through an adaptation of the GoldenGate genotyping assay implemented on a BeadArray platform, the methylation state of 1536 specific CpG sites in 371 genes (one to nine CpG sites per gene) was measured in a single reaction by multiplexed genotyping of 200 ng of bisulfite-treated genomic DNA. The assay was used to obtain a quantitative measure of the methylation level at each CpG site. After validating the assay in cell lines and normal tissues, we analyzed a panel of lung cancer biopsy samples (N = 22) and identified a panel of methylation markers that distinguished lung adenocarcinomas from normal lung tissues with high specificity. These markers were validated in a second sample set (N = 24). These results demonstrate the effectiveness of the method for reliably profiling many CpG sites in parallel for the discovery of informative methylation markers. The technology should prove useful for DNA methylation analyses in large populations, with potential application to the classification and diagnosis of a broad range of cancers and other diseases.

Base Sequence↗

PubMed2005

Deletions within a CA-repeat-rich region of intron 4 of the human SP-B gene affect mRNA splicing.

Length variants within a CA-repeat-rich region of intron 4 of the human SP-B (pulmonary surfactant protein-B) gene are associated with several lung diseases. The hypothesis that SP-B intron 4 affects mRNA splicing was studied. SP-B minigenes containing exons 1-6 with a normal-sized intron 4 (pBi4normal) or intron 4 containing deletions (pBi4del) of 193, 211, 264 or 340 bp were expressed in CHO (Chinese hamster ovary) cells by transient transfection. Two forms of SP-B transcripts, normal and incompletely spliced, were detected. With pBi4normal, normal-sized SP-B mRNA was the predominant form and a very low amount of incompletely spliced mRNA was present, whereas with the pBi4del variants the amount of normal SP-B mRNAs was lower and the amount of incompletely spliced mRNA was relatively high. Reverse transcription-PCR results and sequencing data indicated that the incompletely spliced SP-B RNA contained intron 4 sequence, and this incompletely spliced RNA was also observed in normal lung. Lung cancer tissues with intron 4 deletions exhibited a larger amount of abnormally spliced RNAs compared with normal lung tissue or cancerous tissue with normal-sized intron 4. The results indicate that intron 4 length variants affect SP-B mRNA splicing, and that this may contribute to lung disease.

Animals↗

PubMed2004

Differential allele expression of host defense genes, pulmonary surfactant protein-A and osteopontin, in rat.

Differential allele-specific expression has been observed in several genes involved in immunity. SP-A and OPN play a role in innate host defense. To determine whether SP-A and OPN are subject to differential allele-specific regulation, we investigated their gene or allele-specific expression in various tissues. The results showed: (1) Tissue-specific expression with high levels in lung (SP-A) and kidney (OPN). (2) Differences in allele-specific expression among individuals and tissues. SP-A showed an exclusively balanced biallelic expression (BB) in lung, but both BB and imbalanced biallelic (IB) expression in colon. Allele expression of OPN was more heterogeneous, e.g. in colon BB (22%), IB (64%), and monoallelic expression (MO) (14%). (3) Differential allele-specific expression was observed in all tissues studied (OPN) or in all extrapulmonary tissues (SP-A). (4) Family studies indicated that inheritable factor(s) may be involved in the regulation of allele-specific expression. (5) Analysis of co-expression of gene-specific alleles from double heterozygous rats revealed lack of coordinate allele expression among SP-A, SP-D, and OPN. We conclude that allele-specific expression occurs among genes of innate host defense. This may yet provide another level of regulatory complexity for molecules involved in the first line of defense.

Alleles↗

PubMed2004

Surfactant protein A, an innate immune factor, is expressed in the vaginal mucosa and is present in vaginal lavage fluid.

Surfactant protein A (SP-A), first identified as a component of the lung surfactant system, is now recognized to be an important contributor to host defence mechanisms. SP-A can facilitate phagocytosis by opsonizing bacteria, fungi and viruses, stimulate the oxidative burst by phagocytes and modulate pro-inflammatory cytokine production by phagocytic cells. SP-A can also provide a link between innate and adaptive immune responses by promoting differentiation and chemotaxis of dendritic cells. Because of the obvious relevance of these mechanisms to the host defence and 'gate keeping' functions of the lower genital tract, we examined human vaginal mucosa for SP-A protein and transcripts and analysed vaginal lavage fluid for SP-A. By immunocytochemistry, SP-A was identified in two layers of the vaginal epithelium: the deep intermediate layer (the site of newly differentiated epithelial cells); and the superficial layer (comprising dead epithelial cells), where SP-A is probably extracellular and associated with a glycocalyx. Transcripts of SP-A were identified by Northern blot analysis in RNA isolated from vaginal wall and shown, by sequencing of reverse transcription-polymerase chain reaction products, to be derived from each of the two closely related SP-A genes, SP-A1 and SP-A2. SP-A was identified in vaginal lavage fluid by two-dimensional gel electrophoresis, and confirmed by mass spectrometry. This study provides evidence, for the first time, that SP-A is produced in a squamous epithelium, namely the vaginal mucosa, and has a localization that would allow it to contribute to both the innate and adaptive immune response. The findings support the hypothesis that in the vagina, as in lung, SP-A is an essential component of the host-defence system. A corollary hypothesis is that qualitative and quantitative alterations of normal SP-A may play a role in the pathogenesis of lower genital tract inflammatory conditions.

Adult↗

PubMed2003

Surfactant protein A and B genetic variants predispose to idiopathic pulmonary fibrosis.

Derangement in pulmonary surfactant or its components and alveolar collapse are common findings in idiopathic pulmonary fibrosis (IPF). Surfactant proteins play important roles in innate host defense and normal function of the lung. We examined associations between IPF and genetic polymorphic variants of surfactant proteins, SP-A1, SP-A2, SP-B, SP-C, and SP-D. One SP-A1 (6A(4)) allele and single nucleotide polymorphisms (SNPs) that characterize the 6A(4) allele, and one SP-B (B1580_C) were found with higher frequency ( P</=0.01) in nonsmoker and smoker IPF ( n=84) subgroups, respectively, compared with healthy controls ( n=194). To explore whether a tryptophan (present in 6A(4)) or an arginine (present in other SP-A1 alleles and in all SP-A2 alleles) at amino acid 219 alters protein behavior, two truncated proteins that varied only at amino acid 219 were oxidized by exposure to ozone. Differences in the absorption spectra (310-350 nm) between the two truncated recombinant SP-A proteins were observed both before and after protein oxidation, suggesting allele-specific aggregation differences attributable to amino acid 219. The SP-B SNP B1580_C (odds ratio:7.63; confidence interval:1.64-35.4; P</=0.01), to be a risk factor for IPF smokers, has also been shown to be a risk factor for other pulmonary diseases. The SP-C and SP-D SNPs and SP-B-linked microsatellite markers studied did not associate with IPF. These findings indicate that surfactant protein variants may serve as markers to identify subgroups of patients at risk, and we speculate that these contribute to IPF pathogenesis.

Adult↗

PubMed2002

Heterogeneous allele expression of pulmonary SP-D gene in rat large intestine and other tissues.

Random allele expression has recently been observed for several genes including interleukins and genes of the lymphoid system. We studied the hypothesis that the surfactant protein D (SP-D) gene, an innate host defense molecule, exhibits random allele expression in a tissue-specific manner. SP-D gene expression is tissue specific in the 14 tissues studied. Study of SP-D allelic expression in several tissues revealed a balanced biallelic (BB) in lung, and, in several extrapulmonary tissues, a heterogeneous pattern: BB, imbalanced biallelic (IB), and monoallelic (MO). The results from 103 heterozygous rats showed an expression profile in large intestine of BB (22%), IB (58%), and MO (20%). Among eight families, the percent of BB in siblings varied from 0 to 41%, MO from 0 to 33%, and IB from 49 to 83%. The parent-of-origin does not play a role in SP-D allele-specific expression. However, acquired epigenetic factors, family background, or other factors may contribute to the overall pattern of expression.

Alleles↗