Tracking the progeny of adoptively transferred virus-specific T cells in patients posttransplant using TCR sequencing.

Adoptive cellular therapies with T cells are increasingly used to treat a variety of conditions. For instance, in a recent phase 1/2 trial, we prophylactically administered multivirus-specific T-cell products to protect recipients of T-cell-depleted allogeneic stem cell grafts against viral reactivation. To establish treatment efficacy, it is important to determine the fate of the individual transferred T-cell populations. However, it is difficult to unequivocally distinguish progeny of the transferred T-cell products from recipient- or stem cell graft-derived T cells that survived T-cell depletion during conditioning or stem cell graft manipulation. Using messenger RNA sequencing of the T-cell receptor ß-chains of the individual virus-specific T-cell populations within these T-cell products, we were able to track the multiple clonal virus-specific subpopulations in peripheral blood and distinguish recipient- and stem cell graft-derived virus-specific T cells from the progeny of the infused T-cell products. We observed in vivo expansion of virus-specific T cells that were exclusively derived from the T-cell products with similar kinetics as the expansion of virus-specific T cells that could also be detected before the T-cell product infusion. In addition, we demonstrated persistence of virus-specific T cells derived from the T-cell products in most patients who did not show viral reactivation. This study demonstrates that virus-specific T cells from prophylactically infused multiantigen-specific T-cell products can expand in response to antigen encounter in vivo and even persist in the absence of early viral reactivation.

Iron depletion: an ameliorating factor for sickle cell disease?

We report some observations from our laboratory practice that might be important for the treatment of sickle cell disease (SCD). We describe data from two cases indicating that iron depletion might have a beneficial effect diminishing the formation of HbS in favor of HbF, possibly reducing the severity of the disease. We believe that it would be worthwhile to monitor the course of the disease comparing cases with identical genotypes with and without iron depletion, and we advise to consider chelation therapy to reduce iron overload in patients with SCD.

Fecal continence after rectocele repair: a prospective study.

Combined transvaginal/transanal rectocele repair was performed in series of 89 consecutive women (mean age 55, range 35-81 years) with obstructed defecation due to a rectocele with a depth of more than 3 cm. The impact of this procedure on anal sphincter pressure and continence status was evaluated prospectively. Anorectal manometry was carried out before and after surgery (at 3, 6, 12, and 24 months). The following measurements were performed: maximal anal resting pressure (MARP), maximal anal squeeze pressure (MASP), and rectal sensory perception including first initial sensation, urge to defecate, and maximum tolerable volumes (MTV). The outcome was successful in 71% of patients with respect to symptoms such as the need for straining at defecation, manual assistance, feelings of incomplete evacuation, sense of rectal fullness, constipation, abdominal pain, and the use of laxatives. However, after rectocele repair seven patients experienced deterioration in fecal continence, and dyspareunia developed in 41% of the sexually active patients. Manometric studies revealed a significant decline in mean of 18% of MARP and 16% of MASP. In contrast to MASP, MARP gradually improved during the follow-up period. Distending volumes required for initial sensation and urge to defecate did not change after the procedure. MTV values were significantly lower 3 and 6 months after rectocele repair than those before and 24 months after surgery. MARP and MASP values after surgery did not differ between patients with impaired and those with normal continence. In conclusion, transvaginal/transanal rectocele repair is beneficial for patients with obstructed defecation; however, care should be taken in sexually active patients, and patients at risk of developing fecal incontinence.

Vaccination with experimental feline immunodeficiency virus vaccines, based on autologous infected cells, elicits enhancement of homologous challenge infection.

Cats were vaccinated with fixed autologous feline immunodeficiency virus (FIV)-infected cells in order to present viral proteins to the immune system of individual cats in an MHC-matched fashion. Upon vaccination, a humoral response against Gag was induced. Furthermore, virus-neutralizing antibodies were detected in a Crandell feline kidney cell-based neutralization assay, but not in a neutralization assay based on primary peripheral blood mononuclear cells. Despite the induction of these FIV-specific responses, vaccinated cats were not protected. Instead, accelerated virus replication was found, an observation similar to what previous experiments using other vaccine candidates have shown. Here, the results of the present study are discussed in the light of enhancement of lentivirus infections as a complicating factor in lentivirus vaccine development.

Salmonella typhimurium aroA recombinants and immune-stimulating complexes as vaccine candidates for feline immunodeficiency virus.

Two experimental feline immunodeficiency virus (FIV) vaccines were tested, either alone or in combination, in four groups of cats (A-D). One vaccine (SL3261-FIV) was composed of live attenuated Salmonella typhimurium aroA (SL3261) strains expressing the capsid (Gag) and part of the envelope (Env) proteins of FIV. The other was composed of FIV Gag and Env proteins incorporated into immune-stimulating complexes (iscom-FIV). Cats of group A were immunized four times with SL3261-FIV. Cats of group B were immunized twice with SL3261-FIV and then twice with iscom-FIV. Cats of group C were immunized twice with SL3261 expressing the B subunit of cholera toxin (SL3261-CtxB) and then twice with iscom-FIV. Cats of group D, which served as negative controls, were immunized twice with SL3261-CtxB and then twice with iscom into which the Gag and Env proteins of simian immunodeficiency virus (SIV) had been incorporated (iscom-SIV). Two weeks after the last immunization, all cats were challenged with FIV. At this time, cats immunized with iscom-FIV (groups B and C) showed strong plasma antibody responses to Gag and Env, whilst these responses were weak or undetectable in the cats immunized four times with SL3261-FIV (group A). Seven weeks after FIV challenge, Env-specific antibody responses had increased considerably in cats of all groups except group A. The mean virus loads in the cats of this group proved to be lower than those of the other groups at all time points, indicating partial protection.

Role of defecography in predicting clinical outcome of rectocele repair.

PURPOSE: The aim of this study was to evaluate the role of defecography in predicting clinical outcome of rectocele repair. METHODS: Between January 1988 and July 1994, 74 consecutive patients (median age, 54 (range, 35-81) years) with a rectocele and symptoms of obstructed defecation were studied prospectively. After preoperative evaluation by a standardized questionnaire, physical examination, and defecography, a combined transvaginal/transanal rectocele repair was performed. At follow-up, all patients had defecography. Long-term results were qualified by an independent observer after a median follow-up of 58 (range, 14-89) months as "excellent," "good," or "poor." RESULTS: Rectocele repair was considered excellent in 37 patients and good in 13 patients. Defecography six months after surgery did not show persistent or recurrent rectocele in any of the patients. Size of the rectocele, barium-trapping in the rectocele, internal intussusception, rectal evacuation, and perineal descent did not appear to influence clinical outcome. Radiologic evidence of anismus did not correlate with long-term results of rectocele repair. CONCLUSIONS: Combined transanal/transvaginal repair of rectocele is an efficient therapy in patients with obstructed defecation. Various defecographic parameters (size of rectocele, internal intussusception, rectal evacuation, perineal descent, radiologic signs of anismus) do not appear to influence clinical outcome of surgery. The main value of defecography is the objective demonstration of rectocele and any associated abnormalities such as an enterocele preoperatively and again in objective assessment of the postoperative results.

Enhancement of feline immunodeficiency virus infection after immunization with envelope glycoprotein subunit vaccines.

Cats were immunized three times with different recombinant feline immunodeficiency virus (FIV) candidate vaccines. Recombinant vaccinia virus (rVV)-expressed envelope glycoprotein with (vGR657) or without (vGR657 x 15) the cleavage site and an FIV envelope bacterial fusion protein (beta-Galactosidase-Env) were incorporated into immune-stimulating complexes or adjuvanted with Quil A. Although all immunized cats developed antibodies against the envelope protein, only the cats vaccinated with the rVV-expressed envelope glycoproteins developed antibodies which neutralized FIV infection of Crandell feline kidney cells. These antibodies failed to neutralize infection of thymocytes with a molecularly cloned homologous FIV. After the third immunization the cats were challenged with homologous FIV. Two weeks after challenge the cell-associated viral load proved to be significantly higher in the cats immunized with vGR657 and vGR657 x 15 than in the other cats. The cats immunized with vGR657 and vGR657 x 15 also developed antibodies against the Gag proteins more rapidly than the cats immunized with beta-Galactosidase-Env or the control cats. This suggested that immunization with rVV-expressed glycoprotein of FIV results in enhanced infectivity of FIV. It was shown that the observed enhancement could be transferred to naive cats with plasma collected at the day of challenge.

Interference of imferon in colorimetric assays for iron.

Imferon (Merrell-National), an iron-dextran complex, is widely used in patients with iron deficiency. It is present in the circulation in appreciable amounts for two to three weeks after administration and interferes with all tested colorimetric iron assays, both with and without deproteinization. The amount of the plasma Imferon iron interference depends primarily on the choice of reductant. With dithionite it is essentially 100%. In the presence of ascorbic acid and hydroxylamine, the interference depends also on assay conditions, especially temperature, but also incubation time and pH. The minimum interference in a homogeneous assay was about 3%. The relative amount of interference from hemoglobin iron under the various assay conditions is different from that of Imferon iron. In the presence of a reducing agent, the dextran-iron complex decomposes--instantaneously with dithionite, and gradually with sulfite, ascorbic acid, and hydroxylamine. The freed iron becomes dialyzable, can react with bathophenanthroline, and elutes on a Sephadex G-50 or G-15 column in the same fractions as an ammonium ferrous sulfate.