Cancer immunotherapy using dendritic/tumour-fusion vaccine induces elevation of serum anti-nuclear antibody with better clinical responses.

Dendritic cell (DC) vaccines might induce both anti-tumour immunity and autoimmunity. In this report, we demonstrate elevated levels of anti-nuclear antibody (ANA) in the sera of patients with cancer who had received immunotherapy with a dendritic/tumour-fusion vaccine. Twenty-two patients were treated with DC vaccine of fusion cells composed of autologous DCs and tumour cells (DC/tumour-fusion vaccine), which was generated by treating each cell type with polyethylene glycol. Nine of the 22 patients were treated with both the DC/tumour-fusion vaccine and systemic administration of recombinant human interleukin (rhIL)-12. Serum levels of ANA were examined with an enzyme-linked immunosorbent assay kit. One patient with gastric carcinoma (patient 1, DC/tumour-fusion vaccine alone), one patient with breast cancer (patient 2, DC/tumour-fusion vaccine alone) and one patient with ovarian cancer (patient 3, DC/tumour-fusion vaccine + rhIL-12) showed significant elevations of serum ANA levels during treatment. In patient 1 malignant ascitic effusion resolved and serum levels of tumour markers decreased. Patients 2 and 3 remained in good physical condition during treatment for 24 and 9 months, respectively. Immunoblot analysis indicated antibody responses to autologous tumour cells after vaccination in patient 2. None of the treated patients showed clinical symptoms suggesting autoimmune disease. Patients with elevated serum levels of ANA had significantly longer treatment periods than those without it. Elevated serum levels of ANA after DC/tumour-fusion cell vaccine might be associated with anti-tumour immune response induced by the vaccination.

Cancer immunotherapy by fusions of dendritic and tumour cells and rh-IL-12.

BACKGROUND: Vaccination with fusion cells (FCs) comprising dendritic cells and tumour cells as well as administration of interleukin-12 (IL-12) showed a significant therapeutic effect against established tumours in mouse experimental models. We conducted immunotherapy against various malignant tumours using the FCs and rhIL-12, and investigated the safety and efficacy of the therapy. MATERIALS AND METHODS: Patients' DCs were mixed with autologous irradiated tumour cells and treated with 50% polyethylene glycol to generate FCs. The FCs were inoculated intradermally, and then 30 ng kg(-1) of rhIL-12 was injected at the same sites 2 and 6 days later. This process was carried out as one cycle, and three of these cycles were repeated at 1-week intervals to comprise one course. After completing the course, its safety and therapeutic effects were estimated. RESULTS: The most frequently observed adverse event was fever, observed in 26% of patients in the first cycle. Decrease in white blood cell and an increase in serum ALT were observed in 28% and 25%, respectively. Three out of 12 patients with a malignant brain tumour (25%) achieved a partial response (PR), but other patients with a malignant tumour showed no regression of their tumours. Thirteen out of 16 patients with a brain tumour (81%) showed cutaneous delayed hypersensitivity responses. However, only one of 16 patients (6%) with a malignant tumour other than a brain tumour developed such responses. CONCLUSIONS: Immunotherapy using a FC vaccine and rhIL-12 induced no serious adverse reactions, and provided good therapeutic responses in some of the patients with a brain tumour.

Immunomodulatory effects of selective leucocytapheresis as a new adjunct to interferon-alpha2b plus ribavirin combination therapy: a prospective study in patients with high plasma HCV viraemia.

Efficacy of interferon-alpha2b (IFN) + ribavirin (IFN/RBV) combination in patients with high plasma hepatitis C virus (HCV) is very poor. Dysregulated CD4+ /CD8+ T cells is involved in both impaired cell-mediated immunity and resistance to IFN. Adsorptive granulocytes and monocytes apheresis (GMA) can remove infected leucocytes which are extrahepatic HCV reservoirs and also has been associated with intriguing immunomodulation and increases in CD4+ T cells. Our aim was to see if GMA enhances the efficacy of IFN/RBV. Twenty-four patients, 13 IFN resistant and 11 IFN naive were enrolled. Seventeen were genotype 1b and 7 were 2a or 2b. Mean plasma HCV-RNA was 612.9 (100-850) kIU/mL and alanine aminotransferase, 108 (41-373) U/L. GMA was performed with Adacolumn at one session/day for five consecutive days and IFN/RBV was started within 24 h after the last GMA session. Daily 6 million units of IFN, six times/week for 2 weeks and then three times/week for 22 weeks were given with RBV (600-800 mg/day/patient). Patients were followed for 6 months. GMA was associated with a significant increase in lymphocyte counts, complement activation fragment C3a and falls in tissue necrosis factor-alpha, and IL-8 produced by peripheral blood leucocytes. At week 24, 20 of 24 patients (83%) were HCV negative and by end of follow-up (week 49), the remission was sustained in 14 of 24 patients (58%) including 100% of patients with 2a or 2b. In conclusion, enhanced efficacy of IFN/RBV following GMA might be attributed to a more efficient immune function and a renewed IFN signaling towards HCV.

Association of interleukin-18 gene single-nucleotide polymorphisms with susceptibility to inflammatory bowel disease.

Interleukin-18 (IL-18) is believed to be one of the most important cytokines in the pathogenesis of inflammatory bowel disease (IBD). The aim of the study was to clarify the significance of single-nucleotide polymorphisms (SNPs) at the 5'-end of the IL-18 gene in the development of IBD. DNA was obtained from peripheral blood of 99 patients with ulcerative colitis (UC), 79 patients with Crohn's disease (CD), and 102 healthy controls. All participants were Japanese. SNPs at -656G/T, -607C/A, -137G/C, +113T/G, and +127C/T were determined by means of direct sequencing, and a genetic association with IBD was examined. The frequencies of the G allele at +113 and the T allele at +127 were significantly higher in patients with CD and UC compared with controls. The differences in allelic frequencies were more striking in patients with CD than in patients with UC, and at position +127 than at position +113. The haplotype estimation, according to the E-M algorithm, suggested that TACGT is closely associated with IBD, especially with CD. It was concluded that SNPs at the 5'-end of IL-18 gene might be closely related to the etiology of IBD.

Sisters with atypical Fabry's disease with complete atrioventricular block.

A 56 year old woman with severe right heart failure and complete atrioventricular block was referred to hospital for further examination. Symptoms and signs suggestive of Fabry's disease, such as corneal opacities, acroparaesthesias, hypohidrosis, and angiokeratoma, were not noted. Echocardiography showed a diffuse hypertrophic left ventricular wall and paradoxical movement of the interventricular septum. Cardiac catheterisation showed restrictive-type ventricular dysfunction. Left ventricular endomyocardial biopsy showed central vacuolar degeneration of myocytes with inclusion bodies, which had a concentric lamellar configuration under electron microscopy. This finding is specific for Fabry's disease. The patient's elder sister had experienced an almost identical clinical course and histological findings of myocardial cells on necropsy. In conclusion, two sisters were encountered displaying interesting cases of atypical Fabry's disease. Symptoms began with complete atrioventricular block and histological myocardial findings were specific for Fabry's disease.

Detection of genes expressed in primary colon cancers by in situ hybridisation: overexpression of RACK 1.

AIMS: The isolation of various genes that are expressed in a region specific manner is considered useful for research in molecular pathology. In situ hybridisation (ISH) was used in a screening procedure to isolate these genes efficiently, using colon cancer as a model. METHODS: Suppression subtractive hybridisation (SSH) between colon cancer tissue samples and corresponding non-cancerous tissues was performed. Genes showing high expression in the cancers were selected using macro-DNA array analysis. As a final screening procedure, conventional ISH was performed to isolate genes expressed specifically in colon cancers. RESULTS: Sixty nine clones were selected by SSH and macro-DNA array analyses. These clones were then analysed by ISH to examine their expression patterns. ISH screening revealed that all the clones screened showed more intense signals in colon cancers than in non-cancerous tissues. Among them, RACK 1, which is a protein kinase C receptor and a homologue of the G protein beta subunit, was expressed intensely in colon cancer cells. RACK 1 expression was evaluated in multiple samples by ISH, and the results confirmed that RACK 1 was universally overexpressed in cells of all 11 colon cancers examined. CONCLUSIONS: Many genes, including RACK 1, expressed in colon cancer cells can be isolated efficiently by this method, and their precise expression pattern can be evaluated. These results indicate that ISH is an excellent technique for systemic screening of genes expressed in a region specific manner.

The right ventricular outflow tract as an unusual location for an implantable defibrillator electrode in a patient with arrhythmogenic right ventricular dysplasia.

A 41-year-old woman with arrhythmogenic right ventricular dysplasia (ARVD) underwent the implantation of an implantable cardioverter-defibrillator (ICD), in which the defibrillator electrode was unusually located in the right ventricular (RV) outflow tract. Although fractionated electrograms were demonstrated in the RV apex, which is the usual site for ICD electrodes, normal electrograms were recorded in the RV outflow tract during an electrophysiologic study. An electrode with a screw-in tip was used to fix the implant in the RV outflow tract and obtain successful defibrillation. If normal electrograms are recorded in the RV outflow tract, the site may prove to be an alternative location for an ICD electrode even for ARVD patients.

[Treatment of chronic hepatitis C patients complicated with the other disease].

Because of relatively low efficacy, considerable side effect and high cost, HCV infected patients complicated with the other disease(patients with hematologic dysorders, chronic renal failure on hemodialysis, HIV co-infection and collagen disease) have been excluded from large trials evaluating the efficacy of IFN or in combination with ribabirin. So, little is known about treatment of these patients. We analyzed the medical literature focusing on treatment of HCV infection in these patients, to suggest conclusion about indication based on tolerance and efficacy. In case the other disease is well controlled and long term sevival is expected, treatment should be required based on the evaluation of liver function test including liver pathology for preventing liver disease from developing terminal stage. Further studies are needed to better define HCV therapies in these patients.

FZD4S, a splicing variant of frizzled-4, encodes a soluble-type positive regulator of the WNT signaling pathway.

Frizzled-1 (FZD1)-FZD10 are seven-transmembrane-type WNT receptors, and SFRP1-SFRP5 are soluble-type WNT antagonists. These molecules are encoded by mutually distinct genes. We have previously isolated and characterized the 7.7-kb FZD4 mRNA, encoding a seven-transmembrane receptor with the extracellular cysteine-rich domain (CRD). Here, we have cloned and characterized FZD4S, a splicing variant of the FZD4 gene. FZD4S, corresponding to the 10.0-kb FZD4 mRNA, consisted of exon 1, intron 1, and exon 2 of the FZD4 gene. FZD4S encoded a soluble-type polypeptide with the N-terminal part of CRD, and was expressed in human fetal kidney. Injection of synthetic FZD4S mRNA into the ventral marginal zone of Xenopus embryos at the 4-cell stage did not induce axis duplication by itself, but augmented the axis duplication potential of coinjected Xwnt-8 mRNA. These results indicate that the FZD4 gene gives rise to soluble-type FZD4S as well as seven-transmembrane-type FZD4 due to alternative splicing, and strongly suggest that FZD4S plays a role as a positive regulator of the WNT signaling pathway.

Association of polymorphism in the alcohol dehydrogenase 2 gene with alcohol-induced testicular atrophy.

BACKGROUND: Alcohol abuse can induce testicular atrophy, but it only occurs in some alcoholics. Alcohol dehydrogenase (ADH) is located principally on the Leydig cells. METHODS: To investigate whether genetic polymorphism of alcohol dehydrogenase (ADH) 2 and aldehyde dehydrogenase (ALDH) 2 was related to alcoholic testicular atrophy, we determined restriction fragment-length polymorphisms of the ADH2 and ALDH2 genes in 43 Japanese male alcoholics and 50 healthy subjects. An orchidometer was used to determine the testicular size. RESULTS: Less than 16 ml in testicular size was defined as testicular atrophy. Testicular atrophy was found in 24 (55.8%) cases out of 43 alcoholics. Digestion with MaeIII and MboII after polymerase chain reaction amplification showed that the ADH21 allele frequency was significantly higher in patients with testicular atrophy than in those without testicular atrophy (chi2 = 4.665, p = 0.031), whereas no significant association was observed between testicular atrophy and the ALDH2 gene. CONCLUSIONS: The ADH21 allele may be associated with alcoholic testicular atrophy.

Recent advances in the pathology of alcoholic myopathy.

This article represents the proceedings of a symposium at the 2000 ISBRA Meeting in Yokohama, Japan. The chairs were Victor R. Preedy and Junko Adachi. The presentations were (1) Alcoholic myopathy: Past, present and future, by Timothy J. Peters and Victor R. Preedy; (2) Protein adducts in the type I and II fiber-predominant muscles of the ethanol-fed rat, by Simon Worrall, Seppo Parkkila, and Onni Niemela; (3) Hydroperoxides and changes in alcoholic myopathy, by Junko Adachi, Migiwa Asamo, and Yasuhino Ueno; and (4) A close association between testicular atrophy, muscle atrophy, and the increase in protein catabolism after chronic ethanol administration, by Kunihiko Takeda, Masayoshi Yamauchi, Kazuhiko Sakamoto, Masaru Takagi, Hisato Nakajima, and Gotaro Toda.

Mechanisms of alcoholic liver disease: cytokines.

This article represents the proceedings of a workshop at the 2000 ISBRA Meeting in Yokohama, Japan. The chair was Manuela G. Neuman. The presentations were (1) New aspects of hepatic fibrosis, by D. A. Brenner; (2) Cellular immune response in hepatitis C models, by B. Rehermann; (3) The role of interleukin-10 in acute alcoholic hepatitis, by J. Taieb, S. Chollet-Martin, M. Cohard, J. J. Garaud, and T. Poynard; (4) Cytokine-mediated apoptosis in vitro, by M. G. Neuman; (5) Signaling for apoptosis and repair in vitro, by G. G. Katz, R. G. Cameron, N. H. Shear, and M. G. Neuman; (6) Interferons activate the P42/44 mitogen-activated protein kinase and Janus Kinase signal transducers and activation of transcription (JAK-STAT) signaling pathways in hepatocytes: Differential regulation by acute ethanol via a protein kinase C-dependent mechanism, by B. Gao; (7) Genetic polymorphisms of interleukin-1 in association with the development of Japanese alcoholic liver disease, by M. Takamatsu, M. Yamauchi, M. Ohata, S. Saito, S. Maeyama, T. Uchikoshi, and G. Toda; and (8) Increased levels of macrophage migration inhibitory factor in sera from patients with alcoholic liver diseases, by T. Kumagi, S. M. F. Akbar, M. Abe, K. Michitaka, N. Horiike, and M. Onji.

Histologic heterogeneity and mucin phenotypic expression in early gastric cancer.

Although the major histologic type in small gastric cancers, less than 10 mm in diameter, is differentiated-type adenocarcinoma (D.Ca), the incidence of D.Ca and that of undifferentiated-type adenocarcinoma (UD.Ca) is almost the same in all early gastric cancers. Histologic conversion from D.Ca to UD.Ca has been speculated, however, a detailed examination of this phenomenon has not yet been performed. Three-hundred and 51 early gastric cancers (D.Ca, 150 (42.7%) lesions; UD.Ca, 93 (26.4%) lesions; and mixed differentiated and undifferentiated type (D&UD.Ca), 108 (30.8%) lesions; tumor size less than 10 mm in diameter; 64 lesions, more than 10 mm, 287 lesions) were examined histochemically with paradoxical concanavalin A type III and high-iron diamine-Alcian blue (pH 2.5), and immunohistochemically with antigastric mucin antibody. The associations between tumor size, tumor differentiation and phenotypic expression of mucin were examined. Regardless of the tumor size, mucin phenotypic expression in the mucosal lesions examined was preserved. Of 47 cancers with a gastrointestinal mucin phenotype (GIM type) or a gastric mucin phenotype (GM type) measuring less than 10 mm, 35 (74.5%) consisted of D.Ca and 12 (25.5%) of both D&UD.Ca and UD.Ca, while of 224 GIM or GM type cancers measuring more than 10 mm, 64 (28.6%) consisted of D.Ca and 160 (71.4%) of both D&UD.Ca and UD.Ca. Differences between these two groups were statistically significant (P < 0.001). Of 15 cancers with an intestinal mucin phenotype (IM type) measuring less than 10 mm, 12 (80.0%) consisted of D.Ca and three (20.0%) of both D&UD.Ca and UD.Ca, and of 50 IM type cancers measuring more than 10 mm, 35 (70.0%) consisted of D.Ca and 15 (30.0%) of both D&UD.Ca and UD.Ca. Differences between these two groups were not statistically significant. These findings suggest that small D.Ca showing gastric mucin expression may transform into UD.Ca during the progression of early gastric cancer.

Glycogen storage disease associated with left ventricular aneurysm in an elderly patient.

Glycogen storage disease (GSD) types II, III, IV, and V may be associated with cardiomyopathy, but, with the exception of type III GSD, adult cases are extremely rare. A 62-year-old man was found to have GSD and a concomitant left ventricular aneurysm. He had been comparatively well until the age of 62 years, although he had suffered a cerebral infarction at the age of 35 years. The damage caused by glycogen deposition was strictly confined to the myocardium. Left ventriculography revealed a left ventricular aneurysm in the apex. The serial change on electrocardiogram, as well as the findings of the echocardiogram and of cardiac catheterization, resembled those of the dilated phase of hypertrophic cardiomyopathy. However, a left ventricular endomyocardial biopsy specimen revealed central vacuolar degeneration of myocytes with depositions of glycogen. The GSD type remains unknown in the present patient, because the activity of debranching enzyme (type III) measured from the skeletal muscle specimen was normal, whereas that of acid maltase (type II) was slightly low. It is possible that there is a specific malfunction of the acid maltase of the myocardium in the present patient.

Rapid decrease of plasma HCV RNA in early phase of twice daily administration of 3 MU doses interferon-beta in patients with genotype 1b hepatitis C infection: a multicenter randomized study.

Virological response to interferon (IFN) is poor in patients with plasma levels of HCV RNA higher than 1 Meq/ml and genotype 1b hepatitis C viral infection. In 60 patients, a randomized control study was conducted to compare 3 MU of IFN-beta twice daily for four weeks (group A) and 6 MU once a day for four weeks (group B) followed by a four-week administration of 6 MU once a day. The plasma levels of HCV RNA, determined by an amplicore-monitor method, for patients in group A were significantly lower than those for group B at the fourth and eighth day of IFN administration, and complete virological responses were noted in two patients from group A but none in group B. It is concluded that twice daily administration of 3 MU IFN-beta is more effective than once a day 6 MU in the early phase of IFN therapy.

Alteration of antioxidants during the progression of heart disease in streptozotocin-induced diabetic rats.

Involvement of oxidative stress is implicated in the progression of complication of diabetes mellitus. With respect to heart diseases, we have studied role of oxidative stress/antioxidants using rats treated with streptozotocin to induce diabetes (DM). Hemodynamic and echocardiographic measurements showed thickening of the wall and an increase in the internal dimension of the left ventricle (LV) in DM rats at 8th week. Decrease in diastolic posterior wall velocity and rate of LV pressure change, and increase in LV end diastolic pressures also proved cardiac dysfunction. These changes were further developed in DM rats after 12 weeks. Utilizing rat hearts at 8th and 12th weeks, the following estimations were performed. There was a decrease in the activity of Mn-superoxide dismutase (SOD), suggesting abnormal mitochondrial metabolism of reactive oxygen species. The level of glutathione (GSH) decreased concomitant with a decrease in the expression of gamma-glutamylcysteine synthetase (gamma-GCS). The expression of transforming growth factor-beta1 (TGF-beta1), known as a growth factor and a suppressor of GSH synthesis, elevated in DM rat hearts. Immunohistochemical estimation showed an increase in type IV collagen in DM hearts. Collectively, it was suggested a linkage between mitochondrial damage to generate reactive oxygen species and inactivation of Mn-SOD and elevation of the expression of TGF-beta1 to lead suppression of GSH synthesis and induction of fibrous change for the consequent cardiac dysfunction in DM.

Clinical significance of autoantibody to hepatocyte membrane antigen in type 1 autoimmune hepatitis.

OBJECTIVE: By using HepG2 as flow cytometry target, we have reported that autoantibody to hepatocyte membrane antigen (anti-HMA) was frequently found in autoimmune hepatitis (AIH) patients. In this study, we have examined this autoantibody in relation to clinical features in these patients. METHODS: HepG2 cells were incubated with diluted serum and subsequently with FITC-conjugated antihuman immunoglobulin. The results were expressed as relative fluorescence intensity. The prevalence of anti-HMA was estimated by setting the upper limit of mean +/- 3 SD obtained from healthy subjects. RESULTS: We found that the mean relative fluorescence intensity was 1.67+/-0.5 in AIH with low serum ALT level (group 1 AIH), 4.20+/-1.9 in AIH with high serum ALT level (group 2 AIH), and 1.92+/-0.9 in age-matched chronic hepatitis C virus-positive patients. Their positive rate was 37.5% (three of eight) in group 1 AIH, 95.0% (19 of 20) in group 2 AIH, and 33.3% (four of 12) in chronic hepatitis C patients. In 12 group 2 AIH patients, their mean relative fluorescence intensity was significantly decreased during immunosuppressive therapy. The association between serum ALT level and anti-HMA was confirmed by the facts that a significant direct quantitative relationship existed between these two levels and by serial studies of anti-HMA in four AIH patients. Anti-HMA was also detected in five non-B, non-C hepatitis patients having clinical features resembling those of AIH. CONCLUSIONS: The present results have shown that the anti-HMA was tightly associated with the degree of hepatocyte inflammation and that the measurement of anti-HMA may have some advantage in clinical evaluation of some of non-B, non-C hepatitis patients.