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Neisseria gonorrhoeae enters cultured human mucosal cells following binding of a distinct gonococcal opacity (Opa) outer membrane protein to cell surface proteoglycan receptors. We examined the route of internalization that is activated by Opa-expressing gonococci (strain VP1). Microscopy of infected Chang epithelial cells showed that gonococcal uptake was insensitive to monodansylcadaverine (150 microM), which interferes with clathrin-mediated endocytosis. Similarly, indirect immunofluorescence staining for clathrin in infected cells showed distribution of cellular clathrin unaltered from the distribution in noninfected cells. The microtubule inhibitors colchicine (50 microM) and nocodazole (20 microM) but not the microtubule-stabilizing agent taxol (10 microM) caused a moderate (30 to 50%) reduction in gonococcal entry without affecting bacterial adherence. The most dramatic effects were obtained with the microfilament-disrupting agent cytochalasin D (3 microM), which totally blocked bacterial entry into the cells. Double immunofluorescence staining of gonococci and actin filaments in infected cells demonstrated bacterium-associated accumulations of F-actin as an early signal of bacterial entry. The recruitment of F-actin was transient and disappeared once the bacteria were inside the cells. Cytochalasin D disrupted the actin cytoskeleton architecture but did not prevent the recruitment of F-actin by the bacteria. Adherent, noninvasive gonococcal Opa variants lacked the ability to mobilize F-actin. Recombinant Escherichia coli expressing the gonococcal invasion-promoting Opa of gonococcal strain MS11 (Opa50) adhered to the epithelial cells in an Opa-dependent fashion but was not internalized and did not recruit detectable amounts of F-actin. Coinfection with the E. coli recombinant strain and gonococci resulted in specific entry of the diplococci, despite the presence of large numbers of adherent E. coli cells. Together, our results indicate that Opa-mediated gonococcal entry into Chang cells resembles phagocytosis rather than macropinocytosis reported for Salmonella spp. and sequentially involves gonococcal adherence to the cell surface, Opa-dependent and cytochalasin-insensitive recruitment of F-actin, and cytochalasin D-sensitive bacterial internalization.

Summary. Sequential changes in serum erythropoietin (sEPO) levels were measured by radioimmunoassay in six patients receiving autologous rescue (AR) and 11 patients receiving an allogeneic bone marrow transplant (BMT) for malignant disease. Longitudinal studies showed an inverse relationship between sEPO and haemoglobin levels in the autologous rescue and allogeneic transplant patients throughout the 130 d post-transplant study period. Early post-conditioning EPO responses were normal for the haemoglobin level in both groups, but after day 14 post-transplant, erythropoietin production in response to anaemia became impaired in one autologous rescue patient and eight of the 11 allogeneic transplant patients. There was no clear association between late impairment of sEPO production and conditioning therapy, infection, graft-versus-host disease, immunosuppressive therapy or serum creatinine. Blood transfusion requirements were similar for both groups in the first month after transplantation, but from days 31 to 90 post-transplant, BMT patients required an average of 5.5 units per patient compared with 1 unit per patient for the autologous group. Marrow transplant procedures do not affect early EPO responses but may diminish late responses. The potential value of exogenous rHuEPO in hastening engraftment and decreasing transfusion requirements, particularly for those patients who appear to have impaired EPO responses, remains to be shown by clinical trials.

Eight patients with obstructive sleep apnoea and a normal haemoglobin concentration underwent nocturnal studies during which oxyhaemoglobin saturation was recorded continuously with an ear oximeter and serum erythropoietin concentration was measured hourly by means of a radioimmunoassay. Serum erythropoietin concentrations remained within the normal range throughout the study despite falls in oxyhaemaglobin saturation in individuals to 33-78%. There was no relation between the degree of nocturnal hypoxaemia and serum erythropoietin concentrations. The brief cyclical episodes of hypoxaemia typical of obstructive sleep apnoea may not be a sufficient stimulus for erythropoietin secretion.