Paul Y. Lin

Efficient gene transfer using the human JC virus-like particle that inhibits human colon adenocarcinoma growth in a nude mouse model

The JC virus (JCV) may infect human oligodendrocytes and consequently cause progressive multifocal leukoencephalopathy (PML) in patients with immune deficiency. In addition, the virus has also been detected in other human tissues, including kidney, B lymphocytes, and gastrointestinal tissue. The recombinant major structural protein, VP1, of JCV is able to self-assemble to form a virus-like particle (VLP). It has been shown that the VLP is capable of packaging and delivering exogenous DNA into human cells for gene expression. However, gene transfer is not efficient when using in vitro DNA packaging methods with VLPs. In this study, a novel in vivo DNA packaging method using the JCV VLP was used to obtain high efficiency gene transfer. A reporter gene, the green fluorescence protein, and a suicide gene, the herpes simplex virus thymidine kinase (tk), were encapsidated into VLPs in Escherichia coli. The VLP was used to specifically target human colon carcinoma (COLO-320 HSR) cells in a nude mouse model. Intraperitoneal administration of ganciclovir in the tk-VLP-treated mice greatly reduced tumor volume. These findings suggest that it will be possible to develop the JCV VLP as a gene delivery vector for human colon cancer therapy in the future.

Hepatic tuberculosis: comparison of miliary and local form.

The clinical and pathological features of 22 patients, 11 males and 11 females 17–70 years of age (48.0±16.0 years), with hepatic tuberculosis were reviewed. Five patients had no evidence of extrahepatic tuberculosis (local form), and 17 had the miliary form. The clinical features of the miliary and local forms were similar with pyrexia, abdominal pain, hepatomegaly and body weight loss as the main manifestations. The biochemical findings were also quite similar in reversed albumin and globulin (A/G) ratio (2.9/3.5 vs. 3.2/3.4 g/dl) and disproportionate elevation of alkaline phosphatase (ALP) in comparison with bilirubin values but lower levels of alanine aminotransferase (ALT) (40.4±51.0 vs. 170.8±209.4 U/l; p<0.05) and ALP (208.5±138.9 vs. 389.5±271.1 U/l; p<0.05) in the miliary form. Patients with the local form had higher albumin (3.2±0.8 vs. 2.9±0.7 g/dl), aspartate aminotransferase (AST) (160.4±221.7 vs. 65.9±69.7 U/l), and gamma glutamyl-transpeptidase (γGT) (217.0±144.0 vs. 136.0±92.1 U/l), although the differences were not significant. The histopathological features of the miliary form were also similar to the local form with granuloma, caseation, acid-fast bacilli, fatty change and portal fibrosis as the main findings. The local form revealed more severe signs of hepatocytic damage while the miliary form was more wasting. The results suggest that the miliary and local forms of hepatic tuberculosis had quite similar clinical presentations and pathological features. The biochemical tests suggesting hepatic tuberculosis were reversed A/G ratio and disproportionate elevation of ALP. Bei 22 Patienten mit Lebertuberkulose, 11 Männer und 11 Frauen im Alter von 17–70 Jahren (48,0±16,0 Jahre), wurden die klinischen und pathologischen Befunde ausgewertet. Bei fünf Patienten bestand kein Anhalt für eine extrahepatische Tuberkulose (lokale Form) und bei 17 bestand eine miliare Form. Die klinischen Erscheinungen der miliaren und lokalen Form waren ähnlich mit den Hauptsymptomen Fieber, Bauchschmerzen, Hepatomegalie und Gewichtsverlust. Auch in den biochemischen Befunden bestanden Ähnlichkeiten mit umgekehrter Relation von Albumin und Globulin (A/G) (2,9/3,5 beziehungsweise 3,2/3,4 g/dl) und eine im Vergleich zu den Bilirubinwerten dysproportionale Erhähung der alkalischen Phosphatase (ALP) bei niedrigerem Alaninaminotransferase (ALT)-Spiegel (40,4±51,0 gegenüber 170,8±209,4 E/l; p<0,05) und ALP (208,5±138,9 gegenüber 389,5±271,1 E/l; p<0,05) bei der miliaren Form. Patienten mit der lokalen Form hatten höhere Werte von Albumin (3,2±0,8 gegenüber 2,9±0,7 g/dl), Aspartataminotransferase (AST) (160,4±221,7 gegenüber 65,9±69,7 E/l) und Gamma-Glutamyl-transpeptidase (γGT) (217,0±144,0 gegenüber 136,0±92,1 E/l), doch waren die Unterschiede nicht signifikant. Die histopathologischen Befunde der miliaren und lokalen Form waren ebenfalls ähnlich mit den Hauptbefunden Granulombildung, Verkäsung und Nachweis säurefester Stäbchen, Verfettung und portale Fibrose. Bei der lokalen Form waren stärkere Zeichen der hepatozellulären Schädigung festzustellen, die miliare Form war mit stärkerer Gewichtsabnahme verbunden. Die Ergebnisse der Untersuchung lassen schließen, daß die miliare und lokale Form der Lebertuberkulose mit ähnlichen klinischen und pathologischen Befunden einhergehen. Biochemische Parameter, die auf eine Lebertuberkulose schließen lassen, waren umgekehrter A/G Quotient und dysproportionale Erhöhung der ALP.SummaryThe clinical and pathological features of 22 patients, 11 males and 11 females 17–70 years of age (48.0±16.0 years), with hepatic tuberculosis were reviewed. Five patients had no evidence of extrahepatic tuberculosis (local form), and 17 had the miliary form. The clinical features of the miliary and local forms were similar with pyrexia, abdominal pain, hepatomegaly and body weight loss as the main manifestations. The biochemical findings were also quite similar in reversed albumin and globulin (A/G) ratio (2.9/3.5 vs. 3.2/3.4 g/dl) and disproportionate elevation of alkaline phosphatase (ALP) in comparison with bilirubin values but lower levels of alanine aminotransferase (ALT) (40.4±51.0 vs. 170.8±209.4 U/l; p<0.05) and ALP (208.5±138.9 vs. 389.5±271.1 U/l; p<0.05) in the miliary form. Patients with the local form had higher albumin (3.2±0.8 vs. 2.9±0.7 g/dl), aspartate aminotransferase (AST) (160.4±221.7 vs. 65.9±69.7 U/l), and gamma glutamyl-transpeptidase (γGT) (217.0±144.0 vs. 136.0±92.1 U/l), although the differences were not significant. The histopathological features of the miliary form were also similar to the local form with granuloma, caseation, acid-fast bacilli, fatty change and portal fibrosis as the main findings. The local form revealed more severe signs of hepatocytic damage while the miliary form was more wasting. The results suggest that the miliary and local forms of hepatic tuberculosis had quite similar clinical presentations and pathological features. The biochemical tests suggesting hepatic tuberculosis were reversed A/G ratio and disproportionate elevation of ALP.ZusammenfassungBei 22 Patienten mit Lebertuberkulose, 11 Männer und 11 Frauen im Alter von 17–70 Jahren (48,0±16,0 Jahre), wurden die klinischen und pathologischen Befunde ausgewertet. Bei fünf Patienten bestand kein Anhalt für eine extrahepatische Tuberkulose (lokale Form) und bei 17 bestand eine miliare Form. Die klinischen Erscheinungen der miliaren und lokalen Form waren ähnlich mit den Hauptsymptomen Fieber, Bauchschmerzen, Hepatomegalie und Gewichtsverlust. Auch in den biochemischen Befunden bestanden Ähnlichkeiten mit umgekehrter Relation von Albumin und Globulin (A/G) (2,9/3,5 beziehungsweise 3,2/3,4 g/dl) und eine im Vergleich zu den Bilirubinwerten dysproportionale Erhähung der alkalischen Phosphatase (ALP) bei niedrigerem Alaninaminotransferase (ALT)-Spiegel (40,4±51,0 gegenüber 170,8±209,4 E/l; p<0,05) und ALP (208,5±138,9 gegenüber 389,5±271,1 E/l; p<0,05) bei der miliaren Form. Patienten mit der lokalen Form hatten höhere Werte von Albumin (3,2±0,8 gegenüber 2,9±0,7 g/dl), Aspartataminotransferase (AST) (160,4±221,7 gegenüber 65,9±69,7 E/l) und Gamma-Glutamyl-transpeptidase (γGT) (217,0±144,0 gegenüber 136,0±92,1 E/l), doch waren die Unterschiede nicht signifikant. Die histopathologischen Befunde der miliaren und lokalen Form waren ebenfalls ähnlich mit den Hauptbefunden Granulombildung, Verkäsung und Nachweis säurefester Stäbchen, Verfettung und portale Fibrose. Bei der lokalen Form waren stärkere Zeichen der hepatozellulären Schädigung festzustellen, die miliare Form war mit stärkerer Gewichtsabnahme verbunden. Die Ergebnisse der Untersuchung lassen schließen, daß die miliare und lokale Form der Lebertuberkulose mit ähnlichen klinischen und pathologischen Befunden einhergehen. Biochemische Parameter, die auf eine Lebertuberkulose schließen lassen, waren umgekehrter A/G Quotient und dysproportionale Erhöhung der ALP.

Overexpression of a secretory leukocyte protease inhibitor in human gastric cancer

Complementary DNA microarrays have identified aberrantly expressed genes in patients with gastric cancer. One that encodes secretory leukocyte protease inhibitor (SLPI) is among the aberrantly expressed genes and is associated with metastasis in gastric cancers. We evaluated the potential of SLPI expression as a helpful biomarker for detection of gastric cancer. Tumor tissue and matching noncancerous mucosa were obtained from 60 patients immediately after gastric resection. SLPI expression levels were determined by Northern and Western blot tests and quantitative‐reverse transcriptase‐polymerase chain reaction (Q‐RT‐PCR). Paraffin‐fixed tumor tissues were used for immunohistochemistry study in 119 patients. A consistent result was obtained between all examinations except plasma SLPI. SLPI mRNA transcripts and protein were overexpressed in gastric cancer cells, and the depth of wall invasion was significantly greater in serosa‐invading (T3 and T4) cancers compared to the serosa‐free (T1 and T2) cancers. These enhanced expressions were significantly associated with lymph node metastasis, and were significantly higher in stages III and IV, and higher than those in stages I and II. Five‐year survival of patients with lower expression of the SLPI gene was significantly better than among patients with a higher expression. To better understand the function of SLPI in human gastric cancer cells, isogenic SLPI overexpressing cell lines (AZ521) were prepared. The migratory and invasive abilities were increased 4.4‐fold to 6.9‐fold, or 3.0‐fold to 4.1‐fold, respectively, in SLPI‐overexpressing cell lines. The results point to SLPI as a potential prognostic marker for gastric cancer and its function in cell invasion.

Endoscopic Submucosal Dissection for Early Gastric Cancers : Experience From a New Endoscopic Center in Taiwan

Goal To evaluate the efficacy of endoscopic submucosal dissection (ESD) for early gastric cancers (EGCs) at a new endoscopic center. Background ESD is a novel technique that can facilitate en-bloc resection of EGCs, but seldom reported outside Japan. Study A total of 25 consecutive patients (25 lesions) underwent ESD from June 2004 to March 2006. Patients were divided into 2 groups: group A underwent ESD from June 2004 to May 2005 (introduction stage) and group B from June 2005 to March 2006. The following data were obtained: tumor size, tumor location, operative time, and major complication. Results The complete resection was achieved in 20 lesions (success rate 80%). Four out of 10 lesions from group A were removed by conventional endoscopic mucosal resection (EMR) piecemeally after ESD failure. Conversely, 14 patients from group B (n=15) were resected by ESD en-bloc (success rate 93.3%). One patient with microscopic residual tumor after ESD was further treated by surgical resection. The time required for resection was significantly longer in group A when compared with group B (130.5 min vs. 81.5 min, P<0.05). Postoperative complication rate between the 2 groups were similar. One patient with piecemeal EMR recurred in follow-up, and was further treated successfully by EMR. Conclusions ESD is an ideal method for EGC treatment, but it may result in a risk of complication. The complete resection rate can be improved by endoscopists experience. Sophisticated endoscopic hemostasis and clipping skills are essential prior ESD procedures. Conventional EMR techniques are also obligatory during the beginning period.

Granulomatous hepatitis associated with scrub typhus

A 56 year old patient with scrub typhus infection having unusual presentation of hepatic injury resembling acute hepatitis is described. The clinical features of fever, headache, eschar, lymphadenopathy, lymphocytosis and high Rickettsia tsutsugamushi immunofluorescence titres confirmed the diagnosis of scrub typhus. Acute hepatitis was proven by hepatic biochemical tests and liver biopsy. The patient had a complete recovery soon after antibiotic treatment. The presentation of this case suggests that scrub typhus infection should be included in the list of differential diagnosis of acute hepatitis or granulomatous hepatitis, at least in the Asian Pacific region where scrub typhus still prevails.

Glyoxalase-I is a novel prognosis factor associated with gastric cancer progression.

Glyoxalase I (GLO1), a methylglyoxal detoxification enzyme, is implicated in the progression of human malignancies. The role of GLO1 in gastric cancer development or progression is currently unclear. The expression of GLO1 was determined in primary gastric cancer specimens using quantitative polymerase chain reaction, immunohistochemistry (IHC), and western blotting analyses. GLO1 expression was higher in gastric cancer tissues, compared with that in adjacent noncancerous tissues. Elevated expression of GLO1 was significantly associated with gastric wall invasion, lymph node metastasis, and pathological stage, suggesting a novel role of GLO1 in gastric cancer development and progression. The 5-year survival rate of the lower GLO1 expression groups was significantly greater than that of the higher expression groups (log rank P = 0.0373) in IHC experiments. Over-expression of GLO1 in gastric cancer cell lines increases cell proliferation, migration and invasiveness. Conversely, down-regulation of GLO1 with shRNA led to a marked reduction in the migration and invasion abilities. Our data strongly suggest that high expression of GLO1 in gastric cancer enhances the metastasis ability of tumor cells in vitro and in vivo, and support its efficacy as a potential marker for the detection and prognosis of gastric cancer.

Impact of the Cyclin D1 A870G Polymorphism on Susceptibility to Sporadic Colorectal Cancer in Taiwan

PurposeCyclin D1 is a regulatory protein involved in the cell cycle of both normal and neoplastic cells. Polymorphism of this gene at codon 242 in exon 4 has impacts on risk of the early-age onset in several malignant neoplasms, including colorectal cancer. This investigation was designed to evaluate the effect of cyclin D1 gene polymorphism on the risk of colorectal cancer in Chinese migrants of the Taiwanese population.MethodsWe enrolled 831 primary sporadic colorectal cancer patients as the study group and 1,052 age-gender matched healthy individuals as the control group (1,883 total cases) for present study. Cyclin D1 genotypes (AA, AG, GG) were determined using PCR-RFLP analysis on genomic DNA.ResultsThe frequency of G allele was 39.89 percent and 40.96 percent in the study group and the control group, respectively (P = 0.02). The patients were divided into three age groups for statistical analysis. The younger male patients had a higher frequency of AA/AG genotype compared with the controls (odds ratio, 2.75; 95 percent confidence interval, 1–7.9). The effect of AA/AG genotype on colorectal cancer risk was statistically significant for male patients (odds ratio, 1.34; 95 percent confidence interval, 1.04–1.72), but suchphenomenon was not observed in female patients.ConclusionsOur study suggests that the effect of cyclin D1gene polymorphism on colorectal cancer risk is only observed in males and AA/AG genotype of cyclin D1 gene is associated with a higher risk of colorectal cancer in theyounger patients within the Taiwanese population.

Human JC virus-like particles as a gene delivery vector

Introduction: As a viral gene delivery vector, the recombinant JC virus-like particles (VLPs) can be easily generated in large quantities and at low cost. Exogenous genes of interest can be packaged by the VLP without the involvement of viral genetic material and then delivered into any tissue susceptible to JC virus (JCV) to allow gene transduction. Therefore, it should be possible in the future to develop a gene delivery vector using the human JC VLPs that will allow gene therapy. Areas covered: Development of a gene delivery vector using the polyomavirus VLPs is reviewed in this article. The advantages and disadvantages of using JC VLP for gene delivery are discussed. Expert opinion: Human JC VLPs are readily produced and can be engineered with ease; they allow specific targeting without the presence of any viral genetic material. For therapeutic purposes, gene(s) of interest or other compounds can be packaged into the VLP and delivered to JCV-susceptible cells at high efficiency.

Detection and characterization of mineralo-organic nanoparticles in human kidneys.

Ectopic calcification is associated with various human diseases, including atherosclerosis, cancer, chronic kidney disease, and diabetes mellitus. Although mineral nanoparticles have been detected in calcified blood vessels, the nature and role of these particles in the human body remain unclear. Here we show for the first time that human kidney tissues obtained from end-stage chronic kidney disease or renal cancer patients contain round, multilamellar mineral particles of 50 to 1,500 nm, whereas no particles are observed in healthy controls. The mineral particles are found mainly in the extracellular matrix surrounding the convoluted tubules, collecting ducts and loops of Henle as well as within the cytoplasm of tubule-delineating cells, and consist of polycrystalline calcium phosphate similar to the mineral found in bones and ectopic calcifications. The kidney mineral nanoparticles contain several serum proteins that inhibit ectopic calcification in body fluids, including albumin, fetuin-A, and apolipoprotein A1. Since the mineralo-organic nanoparticles are found not only within calcified deposits but also in areas devoid of microscopic calcifications, our observations indicate that the nanoparticles may represent precursors of calcification and renal stones in humans.

Analysis of the size of DNA packaged by the human JC virus-like particle

Previously, it has been demonstrated that the JC virus-like particle (VLP) is able to package DNA in E. coli and deliver the DNA into human colon cancer cells for gene expression. In this study, the maximum size of DNA packaged by the VLP was determined further. Plasmid DNAs with various sizes were packaged by the VLP in E. coli. Human neuroblastoma cells were then infected with the VLPs containing the various sizes of DNA to allow gene expression. In addition, plasmid DNAs packaged in the VLPs were extracted and retransformed back into E. coli under selection to determine the size of the DNA packaged. The results showed that the JC VLP was able to package plasmid DNA in E. coli up to at least 9.4 kbp in size and this size of DNA could be delivered successfully into human neuroblastoma cells for gene expression. The JC VLP is able to package exogenous DNA up to at least 9.4 kbp in size for gene transduction. These findings will help with the development of gene delivery systems using the JC VLP as the gene delivery vector.