Infection of peripheral mononuclear blood cells by hepatitis C virus.

We investigated the infection of peripheral blood mononuclear cells (PBMNC) by hepatitis C virus (HCV) in 5 patients with HCV-related chronic hepatitis. The presence of HCV-RNA-positive and -negative strands was tested with the polymerase chain reaction (PCR) method. In all subjects, HCV-RNA was shown in PBMNC. In 3 cases, HCV-RNA was shown in the T- and B-cell populations, with viral RNA also present in the monocyte-macrophage fraction of two of these. HCV-RNA-negative stranded molecules, indicative of the viral multiplication, were significantly increased in cells maintained in cultures with PHA/PMA stimulation. The results indicate that HCV infect blood mononuclear cells, thus suggesting that this cellular tropism may play a role in HCV infection.     

Detection of the Arg702Trp, Gly908Arg and Leu1007fsinsC polymorphisms of the NOD2/CARD15 gene by real-time PCR with melting curve analysis.

Crohn's disease is a complex disorder, with multiple genetic traits. A frameshift mutation (Leu1007fsinsC) and two missense mutations (Gly908Arg and Arg702Trp) in the NOD2/CARD15 gene are strongly associated with susceptibility to Crohn's disease. The presence of one of these risk alleles confers a 2- to 4-fold increase in the risk of developing Crohn's disease, and the presence of two mutant alleles increases the risk over 20-fold. To facilitate the analysis of these polymorphisms, we developed three LightCycler assays to detect the missense mutations Arg702Trp and Gly908Arg and the frameshift mutation Leu100fsinsC in the NOD2/ CARD15 gene. All three assays can be run simultaneously on one LightCycler using identical cycling parameters. Analysis of 53 DNAs from Crohn's patients helped to identify carriers at allele frequencies similar to other Caucasian populations. The sequencing of such DNAs confirmed the accuracy of the assays. In conclusion, we present three rapid and robust assays to detect the Arg702Trp, the Gly908Arg and the Leu1007fsinsC ins mutations in the NOD2/CARD15 gene [corrected]     

Dopamine D2 receptor imaging with SPECT: studies in different neuropsychiatric disorders.

The purpose of the present study is to visualize and quantify dopamine D2 receptors in the living human brain using an 123I-labeled ligand and the single photon emission computerized tomography (SPECT) technique. S-(-)-Iodobenzamide [S-(-)-IBZM] has been shown to be a highly selective ligand with high affinity for D2 receptors in experimental studies. Five millicuries (185 MBq) of 123I-labeled S-(-)-IBZM was administered intravenously to 12 control subjects, 22 parkinsonian patients under L-Dopa therapy, 12 parkinsonian patients without L-Dopa, 10 unmedicated patients with Huntington's disease, and 12 patients under different neuroleptics. Data collection with a rotating double-head scintillation camera started 1 h after injection and lasted for 50 min. In a semiquantitative approach, a ratio was calculated between mean counts per pixel in the striatum and a region in the lateral frontal cortex, which was 1.74 +/- 0.10 in the control group. A marked reduction of this ratio was found in patients with Huntington's disease (1.38 +/- 0.12; p = 0.0001), no significant changes in untreated parkinsonian patients (1.67 +/- 0.14), but a reduction in L-Dopa-treated cases (1.59 +/- 0.13; p = 0.0014). A curvilinear relationship was found between total daily dose of neuroleptics and the reduction of this ratio. Estimated receptor blockade under full neuroleptic treatment was 75-80%. S-(-)-IBZM binding was reduced with increasing age (p less than 0.01). Specific binding was reduced markedly when the racemic mixture of IBZM was used, and no specific binding was seen with the R-(+)-isomer, demonstrating the stereoselectivity of IBZM binding.(ABSTRACT TRUNCATED AT 250 WORDS)     

Adeno-associated virus vector-mediated systemic delivery of IFN-beta combined with low-dose cyclophosphamide affects tumor regression in murine neuroblastoma models.

PURPOSE: Type I IFNs (IFN-alpha/beta) have shown significant antitumor activity in preclinical models but limited efficacy and significant toxicity in clinical trials. We hypothesized that the antitumor activity of type I IFNs could be enhanced by chronic, low-dose systemic delivery and sought to test this in murine neuroblastoma models. EXPERIMENTAL DESIGN: Continuous liver-generated expression of human IFN-beta (hINF-beta) was achieved through a gene therapy-mediated approach using adeno-associated virus vectors encoding hIFN-beta (AAV hINF-beta). Orthotopic localized retroperitoneal and disseminated models of neuroblastoma were established using three different xenografts. Immunohistochemical analysis and ELISA were used to evaluate the antiangiogenic effect of therapy. RESULTS: The development of both localized orthotopic (retroperitoneal) and disseminated neuroblastoma was prevented in all mice expressing hINF-beta. Continued growth of established retroperitoneal tumors, treated with AAV hINF-beta as monotherapy, was significantly restricted, and survival for mice with established, disseminated disease was significantly prolonged following administration of AAV hINF-beta. Analysis of treated tumors revealed a significant antiangiogenic effect. Mean intratumoral vessel density was diminished and expression of the angiogenic factors vascular endothelial growth factor and basic fibroblast growth factor were both decreased. Finally, combination therapy in which AAV hIFN-beta was used together with low-dose cyclophosphamide resulted in regression of both established retroperitoneal and disseminated disease. CONCLUSIONS: AAV-mediated delivery of hIFN-beta when used as monotherapy was able to restrict neuroblastoma growth due in part to inhibition of angiogenesis. When used in combination with conventional chemotherapy, AAV hIFN-beta was able to effect complete tumor regression.     

The role of dendritic cells during infection

The skin and the mucosa of the respiratory and gastrointestinal tracts are continuously exposed to microorganisms, but only a limited number of these enter the body and cause disease. To resist microbial infection, the host has developed a multitude of defense mechanisms involving the innate and adaptive immune systems. Dendritic cells (DCs) provide the link between these arms of the immune system. The initiation of an immune response is critically dependent on the activation of DCs, which can discriminate between different classes of microorganisms and elicit tailored antimicrobial immune responses. They have an extraordinary capacity to stimulate naive T cells and initiate primary immune responses. In turn, some pathogens interfere with DC function to block or delay their elimination by the host. Progress in understanding the role of DCs in the host response to microbes is reviewed.