Graft failure in a patient with systemic lupus erythematosus (SLE) treated with high-dose immunosuppression and autologous stem cell rescue.

A 32-year-old female with WHO grade IV, dialysis dependent, lupus nephritis was treated with high-dose immunosuppression and autologous stem cell rescue. Stem cells were mobilized with cyclophosphamide (CY) and G-CSF, and 4.07 x 10(6) CD34+ cells/kg were obtained after CD34+ cell selection using the CellPro column. The preparative regimen consisted of CY, and antithymocyte globulin (ATG), with methylprednisolone. After apparent primary engraftment of neutrophils on day 9, the patient developed recurrent neutropenia on day 19. She showed no evidence of engraftment by day 35, and back-up unmanipulated stem cells were given without effect. Subsequently, she received unmanipulated peripheral stem cells (2 x 10(6) CD34+ cells/kg) from an HLA-identical sibling. The patient remained pancytopenic and expired on day 62 from disseminated fungal infection. An autopsy revealed no evidence of hematopoietic recovery. Progenitor cell assays were performed with the patient's stem cells, which were collected prior to transplantation, and serum collected day 27. Morphologic examination of the patient's cell colonies grown in the presence of her serum revealed abnormal shapes and non-adherent cells. There were significantly fewer BFU-e colonies and a trend toward fewer CFU-GM colonies with the patient's cells and serum compared to normal donor cells. We concluded that a substance present in her serum mediated graft failure and prevented engraftment after additional stem cell infusions.

Sensitivity of testicular germ cells to toxicant-induced apoptosis in gld mice that express a nonfunctional form of Fas ligand.

Germ cell apoptosis in testis is essential for functional spermatogenesis. Recent evidence suggests that the Fas signaling system is critical for the regulation of testicular germ cell apoptosis. To further evaluate the Fas signaling system in testis, we examined the incidence of germ cell apoptosis in gld mice that lack a functional Fas-signaling pathway. gld mice have a small, but significant, increase in testis weight and numbers of spermatid heads per testis compared with wild-type mice. In addition, gld mice have a small increase in the spontaneous incidence of germ cell apoptosis, as indicated by characteristic DNA fragmentation via the terminal deoxynucleotidyl-transferase-mediated deoxy-UTP nick end labeling assay. To test the role of the Fas system in toxicant-induced germ cell apoptosis, mice were exposed to either a Sertoli cell- or germ cell-specific toxicant [mono-(2-ethylhexyl)phthalate (MEHP; 1 g/kg) or 5 Gy radiation, respectively]. These two exposure paradigms induced extensive increases in germ cell apoptosis in wild-type mice. However, exposure of gld mice to MEHP caused only a minimal increase in germ cell apoptosis, whereas they were as sensitive as wild-type mice to radiation exposure. These data indicate that the Fas signaling pathway is 1) involved in regulating the numbers of germ cells in the testis, 2) crucial for the initiation of germ cell apoptosis after MEHP-induced Sertoli cell injury, and 3) differentially active in the cell-specific regulation of germ cell apoptosis that occurs as a consequence of Sertoli cell vs. germ cell injury.

Participation of the Fas-signaling system in the initiation of germ cell apoptosis in young rat testes after exposure to mono-(2-ethylhexyl) phthalate.

The Fas-signaling system is composed of the interacting proteins Fas (CD95/APO-1) and Fas ligand (FasL, CD95L, APO-1L) and is proposed to act in the testis as a paracrine signaling mechanism by which FasL-expressing Sertoli cells initiate apoptosis of Fas-bearing germ cells. Here we describe alterations in the expression of Fas and FasL in the testis after the intimate physical association between Sertoli cells and germ cells is disrupted by exposure to the Sertoli cell toxicant mono-2-(ethylhexyl) phthalate (MEHP). Young, 28-day-old Fisher rats were treated with MEHP (2 g/kg po) and killed 0, 3, 6, and 12 h after exposure. Immunohistochemical analyses revealed a significant increase in the numbers of Fas-positive germ cells as well as increases in the expression of Sertoli cell FasL. Western blot analysis demonstrated a time-dependent increase in the production of the soluble form of FasL after MEHP exposure and suggests that it may participate in triggering apoptosis in germ cells that have lost their intimate association with the Sertoli cells. Measurement of Fas in cytosolic and membrane fractions of testis homogenates by Western blot analysis revealed a significant shift of Fas expression into the membrane fraction after MEHP exposure. Taken together, these observations indicate that the Fas-mediated paracrine signaling mechanism participates in triggering apoptosis of germ cells despite the loss of their close physical association with Sertoli cells. A working model is presented to explain the involvement of the Fas-system in stimulating germ cell apoptosis after MEHP exposure.