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Biomedical subjects

J J Liu

Publications and source records attributed to J J Liu.

At least 37 records · Page 2Linked to original sources
PubMed2001

Proinflammatory chemokine induction in keratocytes and inflammatory cell infiltration into the cornea.

PURPOSE: To determine the effect of interleukin (IL)-1alpha and tumor necrosis factor (TNF)-alpha on cytokine, chemokine, and receptor expression in corneal stromal cells; the effect of corneal scrape injury on monocyte chemotactic and activating factor (MCAF) expression and monocyte-macrophage influx into the stroma; and the effect of MCAF and granulocyte colony-stimulating factor (G-CSF) microinjection on inflammatory cell infiltration into the stroma. METHODS: Gene array technology was used to evaluate changes in cytokine, chemokine, and receptor gene expression in stromal fibroblasts in response to IL-1alpha and TNFalpha. Expression of MCAF mRNA and protein was monitored with an RNase protection assay and Western blot analysis, respectively. Keratocyte MCAF protein expression in the rabbit cornea was detected with immunocytochemistry. After epithelial scrape injury, monocytes-macrophages were detected in rabbit corneas, by immunocytochemistry for monocyte-macrophage antigen. Inflammatory cell infiltration after MCAF and G-CSF microinjection into the stroma of mouse corneas was monitored with hematoxylin and eosin staining. RESULTS: IL-1alpha or TNFalpha upregulated the expression of several proinflammatory chemokines in stromal fibroblasts in culture. These included G-CSF, MCAF, neutrophil-activating peptide (ENA-78), and monocyte-derived neutrophil chemotactic factor (MDNCF). MCAF mRNA upregulation was confirmed by RNase protection assay, and MCAF protein was detected by Western blot analysis. MCAF protein was detected in keratocytes at 4 hours and 24 hours after epithelial injury, but not in keratocytes in the unwounded cornea. Corneal epithelial injury triggered the influx of monocytes-macrophages into the corneal stroma in the rabbit. Microinjection of MCAF and G-CSF into mouse cornea resulted in the influx of monocytes-macrophages and granulocytes, respectively, into the stroma. CONCLUSIONS: Proinflammatory chemokine induction in keratocytes is mediated by IL-1alpha and TNFalpha. The proinflammatory chemokines produced by the keratocytes probably trigger the influx of inflammatory cells into the stroma after epithelial injury associated with corneal surgery, contact lenses, or trauma.

Blotting, Western↗
PubMed2001

[Identification of genes related to cell phenotypic transition by differential display analysis].

To identify the genes that are differentially expressed during the phenotypic transition from vascular adventitial fibroblasts to myofibroblasts, the adventitial fibroblasts were cultured from rat thoracic aorta, and myofibroblasts were obtained by treatment of fibroblasts with TGF-beta1. Differential display PCR (DD-PCR) was used to screen for differentially expressed genes by comparison of mRNA extracted from the two cell populations. Bands upregulated or downregulated on DD gels were excised, reamplified, cloned and sequenced. DD results were verified by quantitative PCR and Northern blot analysis.Antisense oligonucleotide was transfected to study the effect of osteopontin on migration of AF. Differential display showed a significant difference in gene expression profile between the two cell types. A transcript that was downregulated in myofibroblasts showed high DNA sequence homology to part of the gene for NADH dehydrogenase subunit 5. An upregulated transcript showed significant sequence homology to osteopontin gene. Quantitative PCR and Northern blot analysis confirmed the DD results. Among the other differential bands detected, 4 candidate sequences showed no homology to the known genes. The AF numbers of migration were significantly decreased by use of OPN antisense oligonucleotide. This study suggests that the downregulation of gene encoding NADH dehydrogenase subunit 5 and upregulation of osteopontin gene and several other unknown genes may be involved in the phenotypic transition of adventitial fibroblasts to myofibroblats. Inhibition of the expression of OPN may play an important role in the process of vascular remodeling.

Animals↗
PubMed2000

PCD1, a novel gene containing PDZ and LIM domains, is overexpressed in several human cancers.

In an effort to discover novel genes differentially expressed in human pancreatic cancer, we have identified a gene named PCD1 (pancreatic cancer derived) that is up-regulated in pancreatic dysplasia and cancer relative to normal pancreatic ductal epithelium. We cloned the full length (4572 bp) of this gene, which encodes a novel protein of 1064 amino acids containing a PDZ domain and a LIM domain. An alternatively spliced form with a deletion of 30 bp in the coding region was also found. In situ hybridization results showed that PCD1 is highly expressed in a significant percentage of colon, breast, liver, lung, pancreas, stomach, and prostate tumor tissues but is expressed in very few normal tissues. Northern blot hybridization confirmed the overexpression of PCD1 in colon and breast tumor tissues and also showed strong expression of PCD1 in the heart as well as in HeLa cells. Real-time quantitative reverse transcription-PCR verified the overexpression of PCD1 in primary colon tumors or in liver metastases relative to normal colon tissues in five of eight patients. The PCD1 gene maps to human chromosome 13q21.33. Because of its high levels of expression in neoplastic tissues and the presence of both PDZ and LIM domains, we suggest that PCD1 may play an important role in cytoskeletal reorganization during carcinogenesis.

Amino Acid Sequence↗
PubMed2000

Herpes simplex virus infection blocks events in the G1 phase of the cell cycle.

Infection of cells in G1 phase with herpes simplex virus (HSV) prevents their progression into S phase (de Bruyn Kops, A., and Knipe, D. M., 1988, Cell 55, 857-868). We have examined G1-phase events in infected cells to determine whether this effect was the result of inhibition of G1 phase progression or of entry into S phase. We observed that HSV infection decreased pRb phosphorylation and induced a new phosphorylated form of pRb. Furthermore, HSV infection prevented the normal G1 increases in cyclin D1 and D3 protein levels, and blocked the normal G1 appearance of new electrophoretic forms of cdk2 and cdk4. Thus, HSV infection inhibits several events that normally occur in the cell cycle during G1 phase, arguing that the HSV-induced block in the cell cycle occurs in early to mid-G1 phase.

Animals↗
PubMed2000

Increased apoptosis in the heart of genetic hypertension, associated with increased fibroblasts.

OBJECTIVE: The present studies were undertaken to identify apoptosis in cardiomyocytes of genetic hypertension and to study the relationship among apoptosis, aging and blood pressure, and the effect of angiotensin-converting enzyme (ACE) inhibitors on apoptosis. METHODS: Apoptosis in the hearts of spontaneously hypertensive rats (SHR) was identified by electron microscopy (EM) and DNA laddering, and quantified from age 3 weeks to 64 weeks in comparison with normotensive rats (WKY). Fibroblasts and protein products of Bcl-2 and Bax were measured by quantitative immunohistochemistry. SHR were treated with ramipril, an ACE inhibitor. RESULTS: The results showed that: (1) ultrastructural characteristics of apoptosis were observed in cardiomyocytes of SHR, with shrinkage of the cell and condensation of the cytoplasm and chromatin. A DNA ladder was shown; (2) a significant increase in apoptosis in SHR began as early as age 4 weeks and reached a plateau at 16 weeks and maintained at high levels up to 64 weeks. Blood pressure (BP) in SHR started to increase significantly at age 5 weeks; (3) fibroblasts were significantly increased in the heart of SHR; (4) the ratio of Bcl-2/Bax was significantly reduced in SHR; and (6) ramipril effectively reduced apoptosis and fibroblasts, and increased the ratio of Bcl-2/Bax. CONCLUSION: Apoptosis occurs in the cardiomyocytes of genetic hypertension although fibroblasts are increased, and a significant, age-dependent increase in apoptosis is observed. The increase in apoptosis occurs before the difference in blood pressure is detectable. The ACE inhibitor ramipril may be useful for prevention of apoptosis in the heart.

Analysis of Variance↗
PubMed2000

Effects of phospholipids on sphingomyelin hydrolysis induced by intestinal alkaline sphingomyelinase: an in vitro study.

Digestion of dietary sphingomyelin (SM) is catalyzed by intestinal alkaline sphingomyelinase (SMase) and may have important implications in colonic tumorigenesis. Previous studies demonstrated that the digestion and absorption of dietary SM was slow and incomplete and that the colon was exposed to SM and its hydrolytic products including ceramide. In the present work, we studied the influences of glycerophospholipids and hydrolytic products of phosphatidylcholine (PC; i.e., lyso-PC, fatty acid, diacylglycerol, and phosphorylcholine) on SM hydrolysis induced by purified rat intestinal alkaline SMase in the presence of 10 mM taurocholate. It was found that various phospholipids including PC, phosphatidylserine (PS), phosphatidylinositol (PI), phosphatidylethanolamine (PE), and phosphatidic acid (PA) inhibit alkaline SMase activity in a dose-dependent manner, with the degree of inhibition being in the order PA > PS > PI > PC > PE. Similar inhibition was also seen in a buffer of pH 7.4, which is close to the physiologic pH in the middle of the small intestine. When the effects of hydrolytic products of PC were studied, lyso-PC, oleic acid, and 1,2-dioleoyl glycerol also inhibited alkaline SMase activity, whereas phosphorylcholine enhanced SMase activity. However, in the absence of bile salt, acid phospholipids including PA, PS, and PI mildly stimulated alkaline SMase activity whereas PC and PE had no effect. It is concluded that in the presence of bile salts, glycerophospholipids and their hydrolytic products inhibit intestinal alkaline SMase activity. This may contribute to the slow rate of SM digestion in the upper small intestine.

Journal Article↗
PubMed1999

Oligopeptide-repeat expansions modulate 'protein-only' inheritance in yeast.

The yeast [PSI+] element represents a new type of genetic inheritance, in which changes in phenotype are transmitted by a 'protein only' mechanism reminiscent of the 'protein-only' transmission of mammalian prion diseases. The underlying molecular mechanisms for both are poorly understood and it is not clear how similar they might be. Sup35, the [PSI+] protein determinant, and PrP, the mammalian prion determinant, have different functions, different cellular locations and no sequence similarity; however, each contains five imperfect oligopeptide repeats-PQGGYQQYN in Sup35 and PHGGGWGQ in PrP. Repeat expansions in PrP produce spontaneous prion diseases. Here we show that replacing the wild-type SUP35 gene with a repeat-expansion mutation induces new [PSI+] elements, the first mutation of its type among these newly described elements of inheritance. In vitro, fully denatured repeat-expansion peptides can adopt conformations rich in beta-sheets and form higher-order structures much more rapidly than wild-type peptides. Our results provide insight into the nature of the conformational changes underlying protein-based mechanisms of inheritance and suggest a link between this process and those producing neurodegenerative prion diseases in mammals.

Fungal Proteins↗
PubMed1999

Imbalanced expression of functionally different WT1 isoforms may contribute to sporadic unilateral Wilms' tumor.

Functional loss of the product of the Wilm's tumor suppressor gene (wt1) has been identified in subsets of familial Wilms' tumors. Previously, four alternative splice products of WT1 were recognized and each was found to regulate transcription of effector genes differently, suggesting that disruption of the normal ratio of these spliced products will disrupt the normal expression patterns of WT1 effector genes and perhaps lead to Wilms' tumor. In support of these suggestions, we found that four of seven cases of sporadic unilateral Wilms' tumor had striking differences in the ratios of the spliced products of WT1 compared with each other and normal kidney. These data indicate that in addition to structural mutations, alterations in the relative amounts of the mature WT1 isoforms may also be important in the etiology of sporadic Wilms' tumor.

DNA-Binding Proteins↗
PubMed1999

Corneal epithelium-specific mouse keratin K12 promoter.

Keratins are structural proteins expressed by epithelial cells. Approximately 30 different keratin proteins have been identified, each with a specific expression pattern in different epithelial cells. The tissue-specific promoter of several keratin genes have been used to direct the expression of transgenes in animals. Keratin K12 and K3 are expressed in differentiated and stratified corneal epithelium, although the relative expression of each appears to vary between species. We targeted the mouse K12 keratin gene in order to develop a tissue-specific promoter that could be utilized to study the functions of genes of interest expressed in the corneal epithelium. Three 5' truncated fragments of the keratin K12 promoter (1.03, 0.71 and 0.25 Kb) showed higher functional and tissue-specific promoter activity in a human corneal epithelial cell line than other cell lines. The 0.25 Kb K12 promoter fragment was also active in cultured rabbit corneal epithelial cells. Thus, increased expression in corneal epithelial cells directed by fragments of the mouse K12 promoter extended across species lines. The paired box homeotic gene 6 (PAX-6), which is involved in controlling eye development, stimulated the activity of keratin K12 promoter.

Animals↗
PubMed1999

Stromal-epithelial interactions in the cornea.

Stromal-epithelial interactions are key determinants of corneal function. Bi-directional communications occur in a highly coordinated manner between these corneal tissues during normal development, homeostasis, and wound healing. The best characterized stromal to epithelial interactions in the cornea are mediated by the classical paracrine mediators hepatocyte growth factor (HGF) and keratinocyte growth factor (KGF). HGF and KGF are produced by the keratocytes to regulate proliferation, motility, differentiation, and possibly other functions, of epithelial cells. Other cytokines produced by keratocytes may also contribute to these interactions. Epithelial to stromal interactions are mediated by cytokines, such as interleukin-1 (IL-1) and soluble Fas ligand, that are released by corneal epithelial cells in response to injury. Other, yet to be identified, cytokine systems may be released from the unwounded corneal epithelium to regulate keratocyte viability and function. IL-1 appears to be a master regulator of corneal wound healing that modulates functions such as matrix metalloproteinase production, HGF and KGF production, and apoptosis of keratocyte cells following injury. The Fas/Fas ligand system has been shown to contribute to the immune privileged status of the cornea. However, this cytokine-receptor system probably also modulates corneal cell apoptosis following infection by viruses such as herpes simplex and wounding. Pharmacologic control of stromal-epithelial interactions appears to offer the potential to regulate corneal wound healing and, possibly, treat corneal diseases in which these interactions have a central role.

Corneal Stroma↗
PubMed1999

Methanol-related deaths in Ontario.

OBJECTIVE: Methanol poisoning accounts for several deaths annually in the province of Ontario. Our study was aimed at identifying the associated epidemiological factors for fatal outcomes following methanol poisoning in order to develop preventative strategies. METHODS: The records of the Ontario Provincial Coroner's Office were reviewed retrospectively for all poison-related, alcohol-related, and chronic alcohol use-related deaths for the period of January 1, 1986 to December 31, 1991. Age, gender, reason for ingestion (accidental or intentional), and source of methanol for each victim were recorded. RESULTS: There were 43 fatalities during this period, 39 males and 4 females with a mean age of 45 years (range 18-80). Suicide attempts accounted for 21 (49%) cases while the remaining 22 (51%) deaths were classified as accidental. Fourteen (64%) of these 22 patients consumed products labeled as methyl alcohol or wood alcohol as a substitute for ethanol. In 3 cases, the accidental ingestion was the direct result of methanol being improperly stored in containers normally associated with ethanol. The remaining 5 patients were poisoned through the consumption of liquor from illicit sources. CONCLUSIONS: Over half of the methanol-related deaths in Ontario are accidental and potentially preventable. Possible preventative strategies include mandatory product relabeling to eliminate the word alcohol, enhanced public education, and the addition of aversive agents to methanol-containing commercial products.

Accidents↗
PubMed1999

Direct comparison of liposomal doxorubicin with or without polyethylene glycol coating in C-26 tumor-bearing mice: is surface coating with polyethylene glycol beneficial?

Sterically stabilized liposome is characterized by a surface coating of polyethylene glycol (PEG) or other polymers that can reduce opsonization of the liposome by plasma proteins. It has a higher plasma area under the concentration-time curve (AUC), which is believed to correlate with better therapeutic efficacy. However, the presence of large molecules on the liposomal surface may reduce the interactions of liposomes with cells and hinder entry of liposomes into the tumor tissue. Using a stable liposomal system composed of distearoyl phosphatidylcholine/cholesterol, we examined the effect of PEG (Mr 2000) on the pharmacokinetics and on the efficacy of liposomal doxorubicin with C-26 syngeneic tumor model in BALB/c mice. The plasma AUC of liposomal doxorubicin with 6 mol-% PEG-modified distearoyl phosphatidylethanolamine (PEG-DSPE) was approximately twice that of liposomal doxorubicin without PEG at various dosages, regardless of whether the mice were tumor-bearing. Paradoxically, the group of mice treated with liposomal doxorubicin without PEG had higher tumor doxorubicin concentrations. The 72-h tumor AUC was 1.44 times that of liposomal doxorubicin with 6% PEG-DSPE. The tumor-accumulation efficiency (AUC(Tumor)/AUC(Plasma)) of liposomal doxorubicin without PEG was 0.87, and this was more than twice that of the liposomal doxorubicin with 6% PEG-DSPE (0.31). At a dose of 10 mg/kg, although both liposomal groups were better than the free drug group in terms of clinically relevant parameters, including toxicity, tumor shrinkage, and survival, there was no difference between the two liposomal drug groups. In this stable liposome system, surface coating with PEG offered no benefit for liposomal doxorubicin in the C-26 tumor model. To enhance the therapeutic index of liposomal doxorubicin, simply increasing plasma AUC by surface coating with PEG may not be satisfactory.

Animals↗
PubMed1999

The relationship between apoptosis and spontaneous hypertension.

OBJECTIVE: To explore the relationship between apoptosis and spontaneous hypertension, and the effect of angiotensin converting enzyme (ACE) inhibitor on apoptosis. METHODS: Male spontaneously hypertensive rats (SHR) and normotensive control rats (WKY) at different ages were used, meanwhile, the treatment of SHR with ramipril, an inhibitor of ACE was administered orally (1 mg.kg-1.d-1) to SHR from 3 to 10 or from 5 to 10 weeks of age. Apoptosis in cardiomyocytes of SHR was quantified by a maximal labeling (Lmax) method and the characteristic features of apoptosis were identified by electron microscopy (EM), in situ labeling of DNA strand breaks with terminal deoxynucleotidyl transferase mediated dUTP end labeling (TUNEL) and autoradiographic analysis of DNA fragments. RESULTS: The results of a quantitative method showed an age-dependent increase in apoptosis in the cardiac tissues of SHR. A significant increase in DNA breaks occurred as early as 4 weeks and continued to increase up to a plateau at 16 weeks in the cardiac tissue of SHR whereas there was no significant change in apoptosis in WKY up to 64 weeks. Moreover, after the treatment of SHR with ramipril, an inhibitor of angiotensin converting enzyme (ACE), from 3 to 10 or from 5 to 10 weeks of age, the DNA fragmentation as well as blood pressure (BP) was reduced significantly compared with that of untreated SHR (P < 0.01), and similar to that of the control WKY. CONCLUSION: There is a significant increase in the apoptosis of SHR cardiac tissues with increasing age, and ramipril can significantly prevent the increase of apoptosis and in blood pressure, which demonstrates that apoptosis may be involved in the pathogenesis of genetic hypertension. The inhibition of apoptosis as well as hypertension by ACE inhibitors may open a new avenue for developing therapeutic approach for hypertension.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1999

The correlativity analysis of six methods of detecting apoptosis.

The aim of this study was to compare six methods of detecting apoptosis induced by extracellular adenosine triphosphate (ATP) in human leukemic lymphocytes with purinergic P2Z receptors. These methods used were electron microscopy (EM), detection of internucleosomal DNA fragmentation by agarose gel electrophoresis, autoradiographic analysis of DNA fragmentation, in situ labeling of DNA strand breaks with fluorescein dUTP and exogenous terminal deoxynucleotidyl transferase (TUNEL), quantitation of 3'-ends of DNA breaks by labeling with alpha32PdCTP(TdT assay), and quantitation of apoptotic cells with fluorescein-annexin V using flow cytometry (FCA). We found EM and detection of DNA ladder pattern by agarose gel electrophoresis to b specific, but lacking in sensitivity. The combination of autoradiography and gel electrophoresis gave an increase in sensitivity of at least 50 fold although, of all the methods, the TdT assay was shown to be most sensitive. The four methods for quantifying apoptosis-EM, FCA, TUNEL and TdT assay proved to be reliable and gave statistically similar results on apoptotic lymphocytes. These observations indicate it is essential to combine specific, sensitive and quantitative techniques in detecting apoptosis.

Adenosine Triphosphate↗
PubMed1998

A point mutation within exon 5 of the WT1 gene of a sporadic unilateral Wilms' tumor alters gene function.

The Wilms' tumor suppressor gene, wt1, encodes a zinc-finger transcription factor, WT1, that represses transcription of a number of growth-promoting genes and inhibits cell growth. The transcripts of wt1 undergo two alternative splicing events, giving rise to four isoforms of mRNA in constant ratios. The first alternative splice introduces an extra exon 5, which encodes 17 amino acid residues inserted between the transcription regulatory domain and the DNA binding domain of WT1. Previously, we demonstrated that the 17-amino acid domain functioned as a transcription repressor when it was fused with the DNA binding domain of WT1. We have now identified a point mutation within exon 5 of wt1 in a sporadic unilateral Wilms' tumor patient. The mutation changes the last of the 17 amino acids from asparagine to serine. The protein isoform of WT1 carrying this mutation exhibited a 2-3-fold lower transcription-repressing activity than wild-type WT1 in transient cotransfection assays. The mutation also decreased growth-inhibiting activity of WT1 in two osteosarcoma cell lines, U2OS and Saos-2. By diminishing transcription-repressing and growth-inhibiting activities of WT1, this naturally occurring mutation within exon 5 of wt1 may disturb the normal function of the protein and lead to the uncontrolled cell growth characteristic of Wilms' tumor.

Alternative Splicing↗