Regional difference in P-glycoprotein function in rat intestine

It has been reported that inhibition of the P-glycoprotein (P-gp) results in the improved absorption of P-gp substrate in the intestinal tract. In fact, the increased permeability of P-gp substrate across the intestinal epithelium was observed following inhibition of P-gp in in vitro experiments. To develop the formulation containing P-gp inhibitor and P-gp substrate for practical use, it is necessary to know whether the results obtained in the in vitro experiments are reproducible at whole body level. It is also important to find out the regional difference of the P-gp activity in the intestinal tract. In this study, we examined whether verapamil, a specific inhibitor of P-gp, improves the absorption of rhodamine123 (Rho123), a substrate of P-gp, from the jejunum, ileum, and colon of rats using the in situ loop method. The water content in the loop decreased during the experiment, resulting in a significant change of the Rho123 concentration in the loop. Thus, to accurately determine the absorption rate of Rho123, it was necessary to measure the water movement. It was found that there was a regional difference in the water movement, i.e., greatest in colon, followed by ileum. Verapamil did not change the water movement in any intestinal regions. When the concentration of Rho123 in the loop was corrected by water movement, the Rho123 clearance was in the order of ileum (1.15 microL/min/cm), colon (0.83 microL/min/cm) and jejunum (0.47 microL/min/cm). In the presence of verapamil, the Rho123 clearance was significantly increased at jejunum and ileum but not in colon (ileum: 2.08 microL/min/cm, colon: 1.14 microL/min/cm, jejunum: 1.28 microL/min/cm). These results suggest that P-gp inhibits the drug absorption in jejunum and ileum. From these results, it is possible to evaluate the role of P-gp and its regional difference in the in situ experiments. In particular, the inhibition of P-gp results in an increase in absorption of the P-gp substrate limited to jejunum and ileum.     

Dimeric but not monomeric soluble CD40 prolongs allograft survival and generates regulatory T cells that inhibit CTL function

BACKGROUND: We previously reported that adenovirus mediated CD40Ig gene therapy (AdCD40Ig) induced long-term acceptance of fully allogeneic rat cardiac allografts, however, the underlying mechanism has not been fully clarified. To address this we have compared the ability of dimeric and monomeric soluble CD40 to prolong allograft survival in vivo and generate regulatory T cells in vitro. METHODS: The ability of CD40Ig (soluble dimmer, containing an Fc region) or CD40/Myc/His (soluble monomer, lacking an Fc region) therapy to generate CD4CD25 regulatory T cells in vitro and to prevent rejection of rat cardiac allografts (ACI to LEWIS) was compared. Immunoregulatory capacity of regulatory T cells generated was determined by suppression of alloantigen specific proliferation and cytotoxicity. RESULTS: Dimeric soluble CD40Ig did not inhibit CD4 T cell proliferation but rather promoted IL-2 production and the generation of CD4CD25 T cells, which regulated alloantigen-specific cytotoxic T lymphocyte activity. Treatment with either AdCD40Ig or purified soluble CD40Ig prolonged the survival of rat cardiac allografts. In contrast, although monomeric soluble CD40/Myc/His suppressed IL-12 production in a similar manner to that achieved by CD40Ig, it did not augment IL-2 production. Moreover, while CD40/Myc/His also generated CD4CD25 T cells, they did not exhibit regulatory activity and administration of soluble CD40/Myc/His failed to prolong cardiac allograft survival. CONCLUSIONS: These results suggest signaling through CD154 in addition to blocking of CD154-CD40 interaction is important for the immunomodulatory effects of soluble CD40Ig. Taken together, our results provide new insight into the mechanism of immunomodulation by soluble CD40 constructs.     

Health-related quality of life (HRQOL) in Japanese osteoporotic patients and its improvement by elcatonin treatment

Health-related quality of life (HRQOL; QOL hereafter) was evaluated in Japanese osteoporotic patients using three questionnaires; the SF-36 (MOS 36-Item Short-Form Health Survey; generic, profile-type), the EQ-5D (Euro Qol-5 Dimensions; generic, preference-based), and the JOQOL (Japanese Osteoporosis Quality of Life 1999; disease-targeted). The eight subscales and two summary scores of the SF-36 were impaired in these patients even after correction for age and sex. The scores on the EQ-5D and JOQOL correlated well with the subscales of the SF-36 that represent the physical aspects of physical function and bodily pain, which suggests that physical aspects are important determinants of overall QOL status in osteoporotic patients. Although the QOL scores did not correlate with bone mineral density, they were markedly influenced by the presence of vertebral fractures. In particular, the presence of two or more vertebral fractures greatly decreased the QOL scores. We then evaluated the QOL scores before and after treatment. The patients were either given calcium supplementation alone or calcium plus once-weekly elcatonin (Elcitonin, Asahi Kasei Pharma, Tokyo, Japan) injection. Elcatonin treatment markedly improved diverse aspects of the QOL, whereas calcium alone did not. The current data suggest that osteoporosis, especially in the presence of vertebral fracture, is associated with compromised QOL, and therapeutic intervention for osteoporosis should be evaluated in terms of QOL, as well as in terms of increases in bone mineral density and fracture prevention.     

Quantitative identification of sources of dioxin-like polychlorinated biphenyls in sediments by a factor analysis model and a chemical mass balance model combined with Monte Carlo techniques

The major sources of dioxin-like polychlorinated biphenyls (PCBs) in two sediment cores from Tokyo Bay and Lake Shinji (both in Japan) were identified and their source contributions estimated using two receptor models. The first was a nonnegative constrained factor analysis (FA) model, and the second was a nonnegative constrained chemical mass balance model combined with Monte Carlo techniques (CMB-MC) to take into account the variability and uncertainty in both PCB congener profiles of sources and environmental samples. According to the FA model, variations in the concentrations of dioxin-like PCBs in each sediment core were accounted for almost entirely by two factors, which were considered to correspond to Kanechlors (KCs; Japanese PCB products) and incineration. The CMB-MC model investigated the trends of the burdens from four types of KCs and incineration to the concentrations of dioxin-like PCBs in each sediment core. The results for both sediment cores obtained by both models indicated that the burden from KCs increased gradually beginning in the 1950s, peaked around 1970, and declined thereafter, whereas the burden from incineration increased gradually from the 1950s to the early 1990s. The estimated contribution from incineration to the toxic equivalent concentration of dioxin-like PCBs was comparable to that from KCs.     

Gene therapy-mediated CD40L and CD28 co-stimulatory signaling blockade plus transient anti-xenograft antibody suppression induces long-term acceptance of cardiac xenografts

BACKGROUND: The authors have previously demonstrated that inhibition of CD28 and CD40 ligand (CD40L) co-stimulatory signals by adenovirus-mediated cytotoxic T-lymphocyte-associated (CTL) antigen 4 (A4) immunoglobulin (Ig) and CD40Ig gene therapies induces tolerance or long-term acceptance in rat liver and heart allograft transplantation. In this study, the authors examined whether co-stimulation blockade with a brief course treatment of FK779, a novel leflunomide derivative, would be an ideal strategy for controlling xenograft rejection. METHODS: Hamster hearts were transplanted into Lewis rats. Adenovirus vector coding (Ad) CD40Ig, CTLA4Ig, or LacZ gene (1 x 10(9) plaque-forming units) was administered intravenously to recipient rats 2 days before or immediately after xenografting. FK779 (10 mg/kg/day) was administered orally to recipients for 7 days beginning on day -1. Graft survival, graft histology, and xenoreactive antibodies were examined. RESULTS.: Both untreated and AdLacZ-treated control rats rejected cardiac xenografts, with a median survival time (MST) of 3 days. Co-stimulatory blockade alone by AdCTLA4Ig, AdCD40Ig, or both could not overcome such delayed xenograft rejection (DXR) (MST, 3-4 days). Under a short-course FK779 treatment that suppressed T-cell-independent xenoreactive antibodies, administration of AdCD40Ig (MST, 30.5 days) but not AdCTLA4Ig (MST, 9 days) significantly prolonged xenograft survival as compared with the FK779 monotherapy (MST, 7 days). In contrast, DXR and cellular rejection were controlled successfully and all xenografts were accepted for over 100 days when AdCTLA4Ig and AdCD40Ig were administered under FK779 induction therapy. However, chronic rejection was present in all long-term surviving xenografts. CONCLUSIONS.: Gene therapy-based co-stimulation blockade with FK779 induction treatment seems to be an attractive strategy with which to control xenograft rejection.     

Proximal-type epithelioid sarcoma in a 36-year-old man: closer immunoelectron-microscopic resemblance of the tumor cells to epithelial cells than to mesenchymal cells

Proximal-type epithelioid sarcoma (PES) is a rare neoplasm. We report a case of PES that arose in the perineal subcutis of a 36-year-old Japanese man who died within 4 months of the first clinical sign, probably due to massive pulmonary metastases. In the present study, we analyzed the tumor obtained at surgery, immunohistochemically, immunoelectron-microscopically and genetically. Although the tumor cells in the patient expressed both cytokeratin and vimentin immunohistochemically, they showed epithelial characteristics immunoelectron-microscopically because they had tonofilaments constructed of cytokeratin, not vimentin. In addition, the cytokeratins expressed on the tumor were glandular-type keratins. These findings indicate that PES may be a form of carcinoma in soft tissue. To ascertain the possible origin of the tumor, we compared the tumor immunohistochemically with fetal tissues. Although notochord and fetal peritoneal mesothelium were similar to the tumor antigenically, we could not confirm the specific origin of the tumor. Furthermore, the p53-WAF1 pathway did not contribute to tumorigenesis in the patient because the tumor had no mutation in exons 5-8 of the p53 gene and was immunohistochemically positive for WAF1.     

Cytoarchitecture and intercalated disks of the working myocardium and the conduction system in the mammalian heart

Working and specialized cardiac myocytes and their intercalated disks (ID) in the mammalian heart were examined by transmission and scanning electron microscopy. The NaOH/ultrasonication treatment of cardiac tissues resulted in the digestion of collagen fibers and separation of intercellular junctions. Auricular and ventricular myocytes were cylindrical in shape, bifurcated, and connected end-to-end at the ID. The ID in the working myocardium showed a stair-like profile, consisting of steps (plicate segments) and corresponding risers (interplicate segments). The ventricular myocytes had many steps and risers. The steps were filled with numerous finger-like microprojections, including desmosomes, fasciae adherentes, and small gap junctions. The risers showed the smooth surface, including desmosomes and large gap junctions. The cell strands of the sinoatrial node were oriented linearly, while those of the atrioventricular node formed a reticular network. The ID in both nodal cells was underdeveloped, having few microprojections. Myocytes in the His bundle and its branches were arranged in parallel, and Purkinje cell strands formed reticular networks. The ID in the His-Purkinje system was irregular in appearance, and the microprojections were larger in size and smaller in number than those of working myocytes. There were few microprojections in the sheep Purkinje cells. The gap junctions in the conduction system were few or small in size in the nodal tissue, but large in the His-Purkinje system.     

Liposomal surface-loading of water-soluble cationic iron(III) porphyrins as anticancer drugs

A novel design of anticancer drug delivery system, based on an electrostatic binding of negatively charged liposomes and cationic metalloporphyrins under physiological conditions, is reported. A lack of cytotoxicity of the iron(III) porphyrin-loaded liposomes and an efficient generation of a toxic hydroxyl radical (OH*) from a superoxide anion radical (O2-*) through the iron(III)-catalyzed dismutation and the Fenton-like reaction allow for a targeted necrosis of tumor cells where the concentration of O2-* is locally increased as a result of the reduced activity of superoxide dismutase and catalase in these cells.     

Significant liver injury with dual positive IgM antibody to Epstein-Barr virus and cytomegalovirus as a puzzling initial manifestation of infectious mononucleosis

A 35-year-old man was admitted because of significant hepatic dysfunction with mild splenomegaly and intra-abdominal lymphadenopathy of unknown cause. Infectious mononucleosis was suggested by subsequently detected high fever, pharyngotonsillitis and cervical lymphadenopathy, but IgM to Epstein-Barr virus (EBV) and cytomegalovirus (CMV) showed dual positivity. A definite diagnosis of EBV-induced infectious mononucleosis was established 3 months later on the basis of seroconversion to Epstein-Barr nuclear antigen (EBNA)-IgG positivity and reduced CMV-IgM titer with persistently negative CMV-IgG. This case highlights the initial diagnostic difficulties of EBV-induced infectious mononucleosis particularly in older patients, due to concomitant abnormal humoral immunity and unusual initial manifestations such as significant liver injury and extensive intra-abdominal lymphadenopathy.     

Incapacitating lower limb pain syndrome in cord blood stem cell transplant recipients with calcineurin inhibitor

Calcineurin-inhibitor induced pain syndrome (CIPS) is a newly described entity with a characteristic feature of sudden onset of severe lower limb pain, and high levels of cyclosporine or tacrolimus may be involved in the pathogenesis. This syndrome is rarely seen in recipients of hematopoietic stem cell transplantation (HSCT) compared with other organ transplant recipients, however, heightened awareness of this complication after HSCT may be needed for hematologists, as misdiagnosis can result in catastrophic consequences. We report herein two cases of lower limb pain syndrome, with some clinical features resembling CIPS, occurring during the early phase of cord blood stem cell transplantation for hematological malignancy.