Intermittent antibody-based combination therapy removes alloantibodies and achieves indefinite heart transplant survival in presensitized recipients.

BACKGROUND: It is well established that primed/memory T cells play a critical role in heart transplant rejection. This contributes to the challenges faced in the transplant clinic because current treatments that are efficient in controlling naïve T cell alloresponses have limited efficacy on primed T cell responders. METHODS: Fully MHC-mismatched heart transplantation was performed from BALB/c to C57BL/6 mice presensitized with BALB/c splenocytes 14 days pretransplantation. A combination therapy comprising CD70-, CD154-, and CD8-specific antibodies (Abs) was administered at day 0 and 4 posttransplantation with rapamycin on days 0 to 4. RESULTS: The Ab combination therapy extended heart transplant survival in presensitized recipients from median survival time 8 days (MST) to MST 78 days. A decrease in the number of splenic interferon-gamma-secreting cells measured by ELISpot assay was seen in the treated group compared with the untreated controls. However, graft-infiltrating CD8+ and CD4+ T cells persisted despite treatment and the number of intragraft CD4+ T cells increased at day 30 posttransplantation. When an additional "rescue therapy" comprising the same Abs was readministered at days 30, 60, and 90 posttransplantation, T cell infiltration was reduced and indefinite graft survival was observed. Furthermore, rescue therapy resulted in gradual decrease in titer and, by day 90 posttransplantation, the complete loss of the preexisting, donor-specific Abs. CONCLUSION: We conclude that our Ab combination therapy extends allograft survival in presensitized recipients. When combined with intermittent Ab-mediated rescue therapy, this results in indefinite allograft survival and a loss of the preexisting, donor-specific Abs from the circulation.

UNRAVELING THE REGULATION OF NITROGEN ASSIMILATION IN THE MARINE DIATOM THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE): DIURNAL VARIATIONS IN TRANSCRIPT LEVELS FOR FIVE GENES INVOLVED IN NITROGEN ASSIMILATION(1).

We examined the diurnal expression of five genes encoding nitrogen-assimilating enzymes in the marine diatom Thalassiosira pseudonana (Hust.) Hasle et Heimdal following a transition from NH4 (+) - to NO3 (-) -supplemented media. The accumulation of nia transcripts (encoding nitrate reductase, NR) following the transition to NO3 (-) -supplemented media was similar to previously reported changes in NR abundance and activity. Nia mRNA levels varied diurnally, and the diurnal oscillations were abolished when cells were transferred to continuous light. Genes encoding chloroplastic (niiA) and cytosolic (nirB) nitrite reductases were identified in the genome of T. pseudonana. NiiA and nirB transcript levels increased within 2 h following the addition of NO3 (-) and varied diurnally. Patterns of diurnal variation in nia, niiA, and glnII (encoding the chloroplast-localized glutamine synthetase) mRNA abundances were similar. NirB and glnN (encoding the cytosolic-localized glutamine synthetase) mRNA levels also oscillated diurnally; however, the oscillation was out of phase with nia, niiA, and glnII. We propose that NO3 (-) is assimilated into organic molecules in both the chloroplast and cytosol of diatoms and that enzymes encoded by nirB and glnN contribute to the ecologically important dark assimilation of NO3 (-) observed in marine diatoms. As with nia, the diurnal variations in niiA, nirB, glnII, and glnN were abolished when cells were transferred to continuous light. Our results demonstrate that transcript accumulation is not circadian controlled, but, rather, changes in metabolic pools triggered by light:dark (L:D) transitions may be important in regulating the cellular mRNA levels encoding these key nitrogen assimilating enzymes.

Homeostatic proliferation of lymphocytes results in augmented memory-like function and accelerated allograft rejection.

Homeostatic proliferation is a normal physiological process triggered by lymphopenia to maintain a constant level of T cells. It becomes the predominant source of new T cells in adulthood after thymus regression. T cells that have undergone homeostatic proliferation acquire the memory phenotype, cause autoimmune disease, and are resistant to tolerance induction protocols. Transplantation is a rare example in which lymphopenia is deliberately induced for its immunosuppressive effect. However, it is not known whether the homeostatic proliferation that follows will have the opposite effect and accelerate rejection. We show that T cells that have undergone homeostatic proliferation acquire a memory phenotype, spontaneously skews toward the Th1 phenotype, even in the absence of antigenic stimulus. Interestingly, in contrast, the percentage of Foxp3(+) regulatory T cells increased by 28-fold following homeostatic proliferation. Using a mouse life-sustaining kidney transplant model, we showed that T cells that have gone through homeostatic proliferation in lymphopenic hosts transformed chronic rejection to acute rejection of a single MHC class II-mismatched kidney allograft. T cells that have undergone homeostatic proliferation consistently cause reliable rejection even when bona fide memory T cells cannot. These functional changes are long-lasting and not restricted to the acute phase of homeostatic proliferation. Our findings have important implications for tolerance induction or graft-prolonging protocols involving leukocyte depletion such as irradiation bone marrow chimera, T cell-depleting Abs, and lymphopenia induced by infections such as CMV and HIV.