Soft-diet feeding decreases dopamine release and impairs aversion learning in Alzheimer model rats.

To examine the effects of soft-diet feeding on the dopaminergic system in a model rat for Alzheimer's disease (AD), we measured dopamine release in the hippocampus using a microdialysis approach and assessed learning ability and memory using step-through passive avoidance tests. Furthermore, we immunohistochemically examined the ventral tegmental area (VTA), which is the origin of hippocampal dopaminergic fibers using tyrosine hydroxylase (TH), a marker enzyme for the dopaminergic nervous system. Feeding a soft diet decreased dopamine release in the hippocampus and impaired learning ability and memory in AD model rats in comparison with rats fed a hard diet; however, TH-immunopositive profiles in the VTA seemed not to be notably different between rats fed a soft diet and those fed a hard diet. These observations suggest that soft-diet feeding enhances the impairment of learning ability and memory through the decline of dopamine release in the hippocampus in AD rats.

Establishment of HTLV-1 carrier mice by injection with HTLV-1-producing T cells.

We injected with HTLV-1-producing T-cells, MT-2, into C3H/HeJ and Balb/c mice intraperitoneally within 24 hours after birth and at one week old. At 3 months old, HTLV-1 provirus was detected in peripheral blood mononuclear cells in both mice. It was also detected in lymph nodes, thymus, spleen and liver of those mice. The antibody response against HTLV-1 Gag antigen was detected in some of Balb/c mice. These findings suggest that the C3H/HeJ and Balb/c mice were infected with HTLV-1 persistently. The HTLV-1-infected mice should be helpful for studying the pathological state of HTLV-1 carriers and for an establishment of a small animal model of HTLV-1 related diseases including ATL.

Pathogenesis and prevention of HTLV-1-associated diseases.

HTLV-1 is an important factor involved in various diseases including adult T-cell leukemia/lymphoma and HTLV-1 associated myelopathy/tropical spastic paraparesis. Amount of HTLV-1 provirus integrated in human peripheral blood mononuclear cells might be a candidate for a risk factor in the manifestation of HTLV-1 associated diseases. Experimental animal models would be useful to dissect the pathogenesis of HTLV-1 associated diseases. We present rat and mouse models of HTLV-1 infection. Using these animal models, we could clarify the intrauterine transmission of HTLV-1, and have found that both genetic background and HTLV-1 infection are important in pathogenesis of HAM/TSP-like rats. We also discuss the preventive measures of HTLV-1 transmission by use of antisense oligonucleotides.

Inhibition of syncytium formation by antisense oligonucleotide phosphorothioates complementary to tax mRNA of human T-cell leukemia virus type 1 (HTLV-1).

HTLV-1 infection is known as the factor to cause adult T-cell leukemia (ATL). Antisense oligonucleotide phosphorothioates against tax gene and control oligonucleotide phosphorothioates were synthesized. Antisense oligonucleotide was complementary to the region of initiation codon of tax gene. Two control oligonucleotides were tax sense and random. HTLV-1-positive human T-cell line, C91/PL and HTLV-1 non-infected human glioma cell line, U251-MG were co-cultured in the presence of antisense or control oligonucleotides for 24 hours. Oligonucleotides used in this study were not toxic at 10 microM concentration. Antisense oligonucleotide against tax gene inhibited 59% the syncytium formation assay at 10 microM concentration.

Isolation of the poly(ADP-ribose) polymerase-encoding cDNA from Xenopus laevis: phylogenetic conservation of the functional domains.

The complete nucleotide (nt) sequence of the Xenopus laevis poly(ADP-ribose) polymerase (PARP)-encoding cDNA was determined. The putative X. laevis PARP protein consists of 1008 amino acids (aa) with a molecular weight of 113 kDa. X. laevis PARP shares 74, 83, 73, 78 and 42% aa sequence homology with the human, bovine, mouse, chicken and Drosophila melanogaster PARPs, respectively. Comparison of the PARP aa sequences among these species showed conservation of two zinc-finger motifs in the DNA-binding domain, and an NAD-binding motif and a Rossmann fold in the catalytic domain. The first Leu of the putative leucine zipper of D. melanogaster PARP is substituted to Lys in X. laevis PARP. All the Glu residues in the leucine zipper are conserved in these six species.

Age-dependent paraparesis in WKA rats: evaluation of MHC k-haplotype and HTLV-1 infection.

Human T-cell leukemia virus type 1 (HTLV-1) infection is shown to be closely associated with HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Although the occurrence of HAM/TSP was reported to be associated with MHC class II, the mechanism is still unclear. The WKA(RT1k) strain of rats was reported to develop HAM/TSP-like paraparesis after HTLV-1 infection, and was suggested to be an animal model of HAM/TSP. We asked whether MHC k-haplotype is specifically involved in the pathogenesis of paraparesis of WKA(RT1k) rats. We injected the HTLV-1 producing human T cells (MT-2 cells) intravenously into WKA(RT1k) rats and MHC congenic WKA.1L(RT1l) rats which have MHC l-haplotype of LEW rats on the WKA background. Positive antibody response to HTLV-1 antigens and presence of provirus in peripheral blood mononuclear cells confirmed that MT-2 cell-injected rats were infected with HTLV-1. Two of 13 MT-2 cell-injected WKA(RT1k) rats and five of 13 MT-2 cell-injected WKA.1L(RT1l) rats developed HAM/TSP-like hindlimb paraparesis between 16 and 26 months old. Interestingly, three of 14 MT-2 cell-uninjected WKA(RT1k) rats and four of 13 MT-2 cell-uninjected WKA.1L(RT1l) rats showed similar paraparesis between 15 and 26 months old. MHC k-haplotype is not specific to the development of paraparesis in WKA(RT1k) rats. The role of aging, genetic background, HTLV-1 infection and other factors on the development of HAM/TSP-like paraparesis in rats are discussed.

A neuropathological study of paraparetic rats injected with HTLV-I-producing T cells.

In order to clarify the pathogenesis of HTLV-I-associated myelopathy or tropical spastic paraparesis (HAM/TSP), we injected HTLV-I-producing rabbit or human T cells intravenously into WKA and F344 rats. Infection was confirmed from increase in the anti-HTLV-I antibody titer and from the presence of HTLV-I proviral DNA. Only WKA rats developed hindlimb paraparesis 78-124 weeks after the injection. Neuropathological examination of 5 rats showed degeneration of the anterolateral and posterior funiculi as well as the peripheral nerves, and this degeneration was characterized by prominent vacuolation and macrophage infiltration. The myelopathy and neuropathy were grossly similar to those in human HAM/TSP. Although pathological changes of the spinal cord were very mild in 2 paretic rats, and similar lesions were found in the spinal cords and peripheral nerves of 2 control WKA rats, the myelopathy, radiculoneuropathy, or both in the paretic rats showed greater severity than in the controls. The contribution of the aging process to the lesions of the spinal cord and peripheral nerve is discussed. It appears possible that HTLV-I may accelerate the aging process and give rise to paraparesis. The precise role of HTLV-I in the pathogenesis of rat paraparesis remains to be elucidated taking the role of the aging process of the spinal cord and peripheral nerve into account.

A tumour vaccine of fixed tumour fragments in a controlled-release vehicle with cytokines for therapy of hepatoma in mice.

BACKGROUND: Cytokines can be strong potentiators for a tumour vaccine, but they have very short life in vivo when administered as a solution. AIMS: To evaluate the slow release of interleukin 2 from a cytokine-vehicle in vitro and in vivo and to evaluate the anti-tumour activity of a new tumour vaccine in vivo. METHODS: The tumour vaccine was composed of formalin-fixed Hepa 1-6 hepatoma tissue fragments, tuberculin and a lipid based vehicle containing granulocyte-macrophage colony-stimulating factor and interleukin 2. The quantity of interleukin 2 release from the cytokine-vehicle in vitro and in vivo was determined by a proliferation assay with CTLL-2 cell line. Hepa 1-6 hepatoma model system with C57BL/6J mice was used to examine protective and therapeutic anti-tumour effect of the vaccine. RESULTS: Release of interleukin 2 from the cytokine-vehicle lasted 5 days in vitro and 3 days in vivo. The vaccine protected 67% of mice from a Hepa 1-6 cell challenge and had a therapeutic effect by prolonging the life span of mice bearing established Hepa 1-6 tumours of 5 mm in diameter. Of the treated mice, 20% became completely tumour-free. CONCLUSIONS: Formalin-fixed tumour fragments and cytokines in controlled-release vehicle are useful in the rational design of tumour vaccines.

Intrauterine transmission of human T-cell leukemia virus type I in rats.

To analyze intrauterine transmission, MT-2 cells, a human T-cell line producing human T-cell leukemia virus type I (HTLV-I), were injected into eight pregnant F344 rats, and cesarean section was performed at day 23 of pregnancy. HTLV-I provirus was detected by PCR in the liver and spleen taken from one of the eight fetuses. Moreover, 71 offspring were delivered by cesarean section from the remaining seven dams and fostered by seven normal rats. HTLV-I provirus was detected in peripheral blood mononuclear cells in 2 of the 71 offspring 4 weeks after cesarean section. These results indicate for the first time the intrauterine transmission of HTLV-I. To confirm the postnatal transmission, MT-2 cells were injected into a dam within 24 h after delivery, and six offspring were fostered by this dam. HTLV-I provirus was detected in peripheral blood mononuclear cells of all six offspring. This animal model may be useful for analysis and prevention of mother-to-child transmission of HTLV-I.

Influence of diabetes mellitus on left ventricular function in patients undergoing coronary artery bypass grafting.

OBJECTIVES: Left ventricular function was assessed by two-dimensional echocardiography before and one year after coronary artery bypass grafting(CABG) in a series of patients with severe coronary artery disease with diabetes mellitus(DM) and without DM(non-DM). METHODS: Twenty-three patients with DM and 50 patients without DM, all with no previous myocardial infarction, underwent two-dimensional echocardiography before CABG and one year after CABG, in a non-matched study. For a matched study, 31 patients without DM who had almost the same left ventricular function as DM patients at the baseline were selected to and compare the rate of improvement in left ventricular function between the DM group and the "matched" non-DM group. RESULTS: In the non-matched study, patient characteristics were not significantly different between the 2 groups except for the incidence of congestive heart failure within one year before CABG, which was significantly higher in the DM group. Fractional shortening was significantly lower in the DM group at the baseline(p < 0.05) and also one year after CABG(p < 0.0001). Significant improvement in fractional shortening was seen in the non-DM group(p < 0.001), but not in the DM group. The left ventricular end-diastolic diameter(LVDd) was significantly larger in the DM group at the baseline(p < 0.01), and was still significantly larger in the DM group at one year after CABG(p < 0.01). No improvement in LVDd was seen in the DM group. In the matched study, fractional shortening of the non-DM group also showed significant improvement after CABG(p < 0.001). Moreover, the rate of improvement in fractional shortening was higher in the non-DM group than in the DM group(p < 0.05). LVDd tended to be larger in the DM group(p = NS). CONCLUSIONS: Left ventricular dysfunction and left ventricular impaired improvement were seen in the patients with DM, and CABG improved left ventricular function in the patients without DM with poor left ventricular function. These findings indicate that CABG therapy may be inadequate for improving left ventricular function in patients with DM and severe left ventricular dysfunction at the baseline.

Angiotensin-converting enzyme inhibitor therapy affects myocardial fatty acid metabolism after acute myocardial infarction.

BACKGROUND: Angiotensin-converting enzyme (ACE) inhibitor therapy has an early mortality benefit in unselected patients with acute myocardial infarction (AMI). However, the effects of ACE inhibition on myocardial fatty acid metabolism in this patient population have not been studied. We tested the hypothesis that ACE inhibitor therapy improves myocardial fatty acid metabolism and decreases mortality rate in patients after AMI. METHODS: Forty-two patients after first anterior AMI and primary angioplasty were randomly assigned to titrated oral enalapril (n = 24) or placebo therapy (n = 18). Iodine 123-labeled 15-(p-iodophenyl)-3-(R,S)-methylpentadecanoic acid (BMIPP) single photon emission computed tomography imaging was performed an average of 4.8 days after AMI and 1 month after AMI. BMIPP abnormalities were quantified as a severity index by a polar map. RESULTS: There were no significant changes in baseline characteristics, cardiac function, and angiographic findings between patients in the enalapril group and patients in the placebo group. However, BMIPP severity index from acute phase to chronic phase was significantly decreased in the enalapril-treated group (118+/-48 to 82+/-36, P<.05), but not in the placebo group (123+/-65 to 115+/-58, P not significant). CONCLUSION: ACE inhibition therapy improved myocardial fatty acid metabolism and regional left ventricular function in patients after anterior AMI. BMIPP single photon emission computed tomography findings imply that this better outcome may be attributable to an improvement of cellular function with ACE inhibitors.

Isolation of cDNAs encoding the catalytic domain of poly(ADP-ribose) polymerase from Xenopus laevis and cherry salmon using heterologous oligonucleotide consensus sequences.

We have isolated and sequenced cDNAs encoding the catalytic domain of poly(ADP-ribose) polymerase (PARP) from Xenopus laevis and Oncorhyncus masou (cherry salmon). The cDNAs were amplified by polymerase chain reaction using heterologous oligonucleotides corresponding to the conserved sequences of mammalian cDNAs as primers. The deduced amino acid sequences of Xenopus laevis and cherry salmon cDNA showed 84.4% and 75.6% similarities to that of human PARP, respectively. In both species, mRNA for PARP was identified as a single band of 4 kb, and PARP mRNA was abundant in ovary and brain. Thus, mixed oligonucleotide-primed amplification is a useful method in the cloning of cDNAs from different species, and the catalytic domain of PARP is conserved structurally among phylogenetically different species, suggesting an importance of poly(ADP-ribosyl)ation.

Long-term prognosis in achieving a 'stent'like' result from balloon angioplasty: 8 years' clinical outcome.

This study evaluated the long-term prognosis of optimal 'stent-like' results, suboptimal results and failure of balloon angioplasty. The clinical data of 108 patients were examined during 8 years following balloon angioplasty. Based on the angiographic results, the patients were divided into 3 groups: Group A (n=59), <25% residual stenosis (ie, optimal stent-like result); Group B (n=43), 26-50% residual stenosis or large dissection (ie, suboptimal result); and Group C (n=6), >50% residual stenosis or stenosis could not be crossed (ie, failed angioplasty). Restenosis occurred in 20 of 43 patients (46.5%) in Group B, but only in 18 of 59 patients (30.4%) in Group A. Achieving stent-like results following balloon angioplasty significantly reduced the incidence of restenosis. Kaplan-Meier curves at 8 years demonstrated a survival rate without serious cardiac events of 83% in patients with stent-like results compared with 58% in those with suboptimal results and 17% in those with failed balloon angioplasty. In conclusion, the major finding of this study is that achieving stent-like results following balloon angioplasty reduces the incidence of restenosis, and 8-year survival without serious cardiac events after balloon angioplasty is significantly better in patients who have a stent-like result.

A simple and reliable method for the detection and quantitation of the human T-cell leukemia virus type-I provirus in peripheral blood mononuclear cells of seropositive blood donors.

Human T-cell leukemia virus type I (HTLV-I) antibody detection has been widely used to screen HTLV-I carriers. Sometimes, however, it gives false positive or negative results. A demonstration of the HTLV-I provirus from patients' peripheral blood mononuclear cells (PBMC) should, therefore, give the crucial evidence for them being HTLV-I carriers. We established a simple and reliable method using the polymerase chain reaction (PCR) to detect one molecule of HTLV-I provirus in 100 x 10(3) PBMC, during which internal control primers for the human beta-globin gene were also employed in the same reaction tube to check the success of the amplification reaction. We can thus easily avoid any false negative judgement and quantitate the HTLV-I provirus in PBMC simply by diluting the sample before PCR. One ml blood was enough for ten or more determinations by PCR. Analysis of seropositive blood from donors demonstrated a wide range for the number of HTLV-I provirus in PBMC. The method could conveniently be used for quantitating HTLV-I proviruses and following up HTLV-I carriers to study the pathophysiology and mode of HTLV-I transmission.

Integration of HTLV-1 provirus into mouse transforming growth factor-alpha gene.

Human T cell leukemia virus type 1 (HTLV-1) provirus integration was investigated in mice inoculated with MT-2 cells, a HTLV-1 producing human T-cell line. From spleen DNA derived from a MT-2 cell-injected mouse, we cloned a HTLV-1 integration site by ligation-mediated PCR. The nucleotide sequence showed that HTLV-1 provirus was integrated into an intron of the mouse transforming growth factor-alpha gene in the reverse orientation. This result provides for the first time molecular evidence that mice can be infected with HTLV-1.

Brugada syndrome characterized by the appearance of J waves.

We describe a patient with Brugada syndrome. The ST-segment elevation in precordial leads was revealed during admission, but the appearance of J waves was characteristic before ventricular fibrillation (VF), rather than ST-segment elevation. J waves have been reported to be associated with the presence of an Ito-mediated prominent action potential notch in the epicardium. It is considered that one of the mechanisms of this VF is due to heterogeneous distribution of the refractory period according to changes in K+ channels including Ito.

Vertical transmission of HTLV-1 in HTLV-1 carrier rat.

A female F344 rat was injected with 2.4 x 10(6) MT-2 cells intravenously at 3 and 4 weeks old, and was mated with a non-infected male rat at the 17th week after injection, when the dam rat contained HTLV-1 provirus in the peripheral blood mononuclear cells as determined by polymerase chain reaction. HTLV-1 provirus was detected in at least 2 out of 9 offspring. This system should be useful for studies on the routes and prevention of vertical transmission and on elimination of once-transmitted HTLV-1.

Infection of rats with HTLV-1: a small-animal model for HTLV-1 carriers.

A human T-cell line producing human T-cell leukemia virus type I (HTLV-I), MT-2, was injected intravenously into female F344 rats aged 5 weeks to make HTLV-I carrier rats. Antibody against HTLV-I was detected at the 5th week after MT-2 injection, and its titer reached a high plateau which continued from the 15th to the 27th week. The antibodies were against p19, p24, p28 and p53 of HTLV-I antigens from MT-2 cells. The gag, pX and LTR nucleotide sequences of HTLV-I provirus were demonstrated by using polymerase chain reaction (PCR) in the peripheral-blood mononuclear cells of 3 rats at the 44th week and 2 at the 66th to 68th week out of 8 F344 rats injected with MT-2 cells. Quantification of the HTLV-I proviral sequence revealed that 30 to 60 molecules were present in 10(5) peripheral-blood mononuclear cells, indicating that the rats were chronically infected with HTLV-I. HTLV-I-infected rats could serve as a small-animal model for studying the pathophysiological state of HTLV-I carriers and also that of HTLV-I infection on various HTLV-I-related diseases, including adult T-cell leukemia and HTLV-I-associated myelopathy.

Inhibition of human T-cell leukemia virus type 1 replication by antisense env oligodeoxynucleotide.

Human T-cell leukemia virus type 1 (HTLV-1) infection is associated with adult T-cell leukemia and HTLV-associated myelopathy/tropical spastic paraparesis. Inhibition of HTLV-1 transmission is important to prevent the above HTLV-1-associated diseases. We used the antisense oligodeoxynucleotides (oligos) complementary to the first splice junction, rex responsive site, gag, env, tax, rex, and p21 and evaluated the effects on the syncytium formation between HTLV-1 producing human T-cell line, C9/PL cells, and HTLV-1-uninfected human glioma cell line, U251-MG cells. The syncytium formation was significantly inhibited the virion production assayed by antisense oligos to env, tax, gag, p21, and rex, with antisense oligo to env being the most inhibitory. Antisense oligos to env and tax also inhibited reverse transcriptase activity. Antisense oligo to env may have a potential as a preventive measure of HTLV-1 replication and transmission in vivo.

High incidence of HAM/TSP-like symptoms in WKA rats after administration of human T-cell leukemia virus type 1-producing cells.

We demonstrate a significantly high incidence of human T-cell leukemia virus type 1 (HTLV-1)-associated myelopathy (HAM)-or tropical spastic paraparesis (TSP)-like symptoms in WKA rats after injection with HTLV-1-producing MT-2 cells, while no symptoms were observed in F344 rats injected with MT-2 cells or in control WKA rats. Five of the eight (63%) WKA rats injected with MT-2 cells showed HAM/TSP-like paraparesis at 105 weeks of age, but none of seven MT-2-injected F344 rats or eight control WKA rats showed symptoms. This high incidence of HAM/TSP-like symptoms in WKA rats was statistically significant (P < 0.05). Six of the eight (75%) WKA rats injected with MT-2 cells showed HAM/TSP-like paraparesis at 108 weeks of age. HAM/TSP-like symptoms were also observed in one of the two WKA rats injected with HTLV-1-producing Ra-1 cells at 128 weeks of age. HTLV-1 provirus was detected in peripheral blood mononuclear cells in both WKA and F344 rats. The provirus was detected in the spinal cords of the HAM/TSP-like WKA rats that had severe neuropathological changes. WKA and F344 rats showed no significant difference in antibody response against HTLV-1 Gag antigen. However, the antibody response against the C-terminal half of gp46 HTLV-1 envelope protein was lower in WKA rats than in F344 rats. Pathological analysis of the HAM/TSP-like rats showed degeneration of the white matter of the spinal cord and peripheral nerves. These findings suggest that both the genetic background of the host and HTLV-1 infection are important in neuropathogenesis of HAM/TSP-like paraparesis in rats.