Dynamics of HIV replication in lymphocytes and the efficacy of protease inhibitors

Using a system that allows transfection of resting peripheral blood lymphocytes (PBLs) two questions were addressed: the kinetics of HIV replication from the state of proviral latency, and the impact of different parameters on the efficacy of protease inhibitors to control HIV replication. PBLs were transfected with an infectious full length HIV-DNA harboring a luciferase reporter gene and activated thereafter. Ritonavir was added at different times at doses ranging from to 0.06 to 1 microM. Viral expression was assessed by quantifying luciferase activity in cell extracts and levels of p24 HIV antigen in culture supernatants. After transfection and cell activation, intracellular expression of HIV proteins, as assessed by luciferase detection, occurred within 2 hr. HIV-gag p24 antigen was detected in culture supernatants between 6 and 8 hr post-activation. Ritonavir was effective in blocking viral replication when given within 4 hr following HIV reactivation, but a delay in ritonavir administration or breaches in ritonavir levels after 6 hr from transfection resulted in viral escape. HIV reactivation from proviral latency in PBLs is an extremely rapid process, faster than estimated from previous models. These data stress the need for maintaining effective antiretroviral concentrations to block completely viral replication.     

Long-term protective immune response elicited by vaccination with an expression genomic library of Toxoplasma gondii

Immunization of BALB/c mice with an expression genomic library of Toxoplasma gondii induces a Th1-type immune response, with recognition of several T. gondii proteins (21 to 117 kDa) and long-term protective immunity against a lethal challenge. These results support further investigations to achieve a multicomponent anti-T. gondii DNA vaccine.     

Evidence of a diverse T cell receptor repertoire for acetylcholine receptor,the autoantigen of myasthenia gravis

We utilized two methods to look for T cell clonal expansions in myasthenia gravis (MG). We analyzed TCRBV CDR3 length polymorphism (spectratyping) to look for evidence of clonal expansion of CD4 or CD8 T cells directly from peripheral blood of MG patients. No statistically significant differences were found between the diversity of TCR repertoires in MG patients compared to normal control individuals when analyzed as groups. Rare oligoclonal expansions were detected in some individual MG patients but the significance of these findings is unclear. Next, we analyzed a panel of T cell hybridomas from acetylcholine receptor (AChR) immunized, MG-susceptible HLA-DR3 transgenic mice. The epitope specificity, TCRBV gene usage and CDR3 sequences of these hybridomas were highly diverse. We conclude there is only limited evidence for restricted TCR repertoire usage in human MG and suggest this may be due to the inability of HLA-DR molecules to select for restricted TCR recognition of AChR epitopes.     

Pregnancy with frozen-thawed and fresh testicular biopsy after motile and immotile sperm microinjection, using the mechanical touch technique to assess viability

BACKGROUND: There are few reports of pregnancy using immotile sperm, and none using a purely mechanical assessment of viability. METHODS: In this pilot study, we retrospectively analysed 66 cycles in 61 patients with determinant male factor, recording rates of fertilization, implantation, normal pregnancy and take-home babies achieved with ICSI. Sperm selection was based on morphologically normal appearance under the inverted microscope. Viability of immotile spermatozoa was assessed by the mechanical touch technique to observe tail flexibility and tail shape recovery. RESULTS: Of 17 ICSI cycles using frozen-thawed testicular sperm, six microinjected with immotile and 11 with motile sperm, we achieved fertilization rates of 65.7 and 74.3%, respectively, and five pregnancies (two and three, respectively). Of 49 ICSI cycles using fresh testicular sperm, 10 microinjected with immotile and 39 with motile sperm, we achieved fertilization rates of 73.4 and 64.4%, respectively, and 12 pregnancies (three and nine, respectively). CONCLUSIONS: Immotile (fresh and frozen-thawed) testicular sperm of normal morphological appearance can be used to achieve clinical pregnancy with ICSI. Our results strongly suggest that immotile sperm viability can be assessed by the mechanical touch technique.     

Meningeal carcinomatosis in bladder tumor

We report a strange case of a bladder whose first metastasic manifestation, after two years of the diagnosis, was a peripheric polyneuropathia. This patient was treated with immunotherapy with BCG for superficial carcinoma of the bladder during one year. Gradually central neurological symptoms appeared and the patient died one month later. A meningeal carcinomatosis was identify as the cause. No bone metastases existed, which is the most frequent way of tumours extension towards leptomeninges. We argue about the way to arrive at meninges.