Enhanced transduction of mouse bone marrow-derived dendritic cells by repetitive infection with self-complementary adeno-associated virus 6 combined with immunostimulatory ligands.

The potential of adeno-associated virus (AAV)-based vectors in human gene therapy is being explored for several diseases. Although sustained transgene expression and low vector-associated cellular immunity are attractive features of recombinant (r) AAV, the wider application of rAAV vectors encapsidated in serotype 2 capsid is hampered by poor transduction efficiency in many target tissues. These include ex vivo-generated dendritic cells (DC), which have demonstrated promising immunotherapeutic activity. We report here that efficient transduction of mouse bone marrow-derived DC can be achieved with self-complementary (sc) rAAV encapsidated in serotype 6 capsid. Sequential exposure of DC precursor cultures to IL-4 and GM-CSF with sc rAAV6 encoding the human tumor antigen, carcinoembryonic antigen (CEA), for 7 days followed by activation with CpG oligodeoxynucleotides (ODN) and anti-mouse CD40 antibody resulted in highly efficient transduction of DC. DC surface markers as determined by flow cytometry analysis of sc rAAV6-transduced DC were comparable to nontransduced DC. Efficiency of vector transduction and transgene expression were confirmed by immunostaining and real-time PCR. Microarray analysis of RNA from CpG ODN and CD40 antibody stimulated sc AAV6-transduced DC revealed upregulation of transcription factors and cytokines involved in immune activation and downregulation of inhibitory factors, suggesting a possible role of transcriptional activation in the observed effect. The adoptive transfer into syngeneic mice of the ex vivo-transduced and activated DC resulted in the development of CEA-specific antibody and T-helper 1-associated immune responses. Immunized mice also developed antibody to AAV6 capsid protein, which did not crossreact with AAV2 capsid protein. These studies demonstrate the potential utility of sc rAAV serotype 6-based vectors in transduction of DC for genetic vaccination approaches.

Evaluation of severe reactions to sweat bee stings.

Thirteen patients with severe reactions to sweat bee stings were evaluated. Eleven patients had had systemic reactions and two, large local reactions. Skin testing and RAST with venom of other Hymenoptera and whole body extract of sweat bees disclosed negative reactions to both skin tests and RAST of all other insects tested in eight, while three showed sensitivity to honey bee. Only one patient had a positive RAST reaction to whole body extract of sweat bee. Whole body extract of sweat bee causes some irritant reaction when skin testing controls and is not a reliable diagnostic agent even though patients were more reactive than controls. Hypersensitivity reactions to sweat bee stings are distinct and not associated with reactions to other stinging insects. Currently, no reliable antigen is commercially available for diagnosing or treating this condition. Sweat bee venom or other venom antigens might be beneficial.

Immunoglobulins and hyposensitization for allergy.

The premise that protracted stimulation of the human immune system might be involved in the induction of malignant immunocytopathies is scrutinized. Studies of the serum immunoglobulins of 313 patients who had been hyposensitized for allergy are compared with those of 138 nonallergic and 189 allergic individuals not subjected to this therapy. Critical historical data in all groups are also evaluated. The investigators concluded that, as far as the parameters studied, there were significant differences in the mean total immunoglobulin G levels in allergic hyposensitized persons compared with the non-allergic control group. It involved a depression of the serum IgG concentration and related to the length of the desensitization program and to whether it was continuous or interrupted. One evolving monoclonal immunocytopathy, etiology of which has not been established, was encountered during the period of study.

The round nipple shadow.

It is usually easy to distinguish the round nipple shadow from a lesion, especially by means of repeat radiographs with nipple markers. However, the radiologist must occasionally interpret a round density in the lower chest on the basis of a single radiograph or series of radiographs. Fortunately, a nipple shadow usually has a well-defined outer margin and a poorly defined inner margin. Differentiation of a nipple shadow from an extrapleural or pleural lesion is described. Attention to these findings may prevent needless additional study and patient apprehension.

Aneurysms secondary to pancreatitis.

In a review of arteriograms of 72 unselected consecutive cases of pancreatitis, seven patients were found to have arterial aneurysms involving branches of the peripancreatic vessels. During the same period, arteriograms of 84 cases of carcinoma of the pancreas were reviewed and no aneurysms of any of these vessels were found. The demonstration of aneurysms of the peripancreatic arteries in pancreatitis is an important differential feature from carcinoma of the pancreas. Both carcinoma of the pancreas and chronic pancreatitis can cause encasement of the arterial vessels and obstruction of the splenic or the superior mesenteric vein, therefore resulting in a similar angiographic appearance. Thus an aneurysm seen in such a patient is a helpful distinguishing feature. In addition, these aneurysms are an important source of hemorrhage and mortality in pancreatitis.