In vitro generation of adherent mononuclear suppressor cells to Toxoplasma antigen.

Adherent mononuclear cells have been found to suppress the lymphocyte proliferation, of T lymphocytes of patients with various chronic infections, to pathogen-specific antigens. To explore mechanisms involved in the generation of these suppressor cells, we established an in vitro method for the generation of suppressor-adherent mononuclear cells. Adherent mononuclear cells separated from mononuclear cells from subjects with serological evidence of chronic Toxoplasma infection could be induced, by preincubation with Toxoplasma antigen for 8 days, to suppress the proliferative response to autologous mononuclear cells to Toxoplasma antigen (TA) (mean suppression = 47%) and tetanus toxoid (TT) (mean suppression = 39%) compared to the proliferative response of autologous mononuclear cells co-cultured with no antigen. When adherent cells were removed after 1-day culture there was no significant suppression of the lympho-proliferative response to TA or TT. Induction of the adherent suppressor cell depended on the presence of CD4-positive T cells and not CD8-positive T cells. Adherent suppressor cells acted directly on the proliferative response of CD4 cells to antigen. The adherent cells contained 90 +/- 5% esterase-positive cells. In cell-mixing experiments, equal numbers of CD8-positive T cells pretreated in a similar manner did not have a suppressive effect. However, pretreated CD4-positive cells did have a suppressive effect at higher concentrations of cells than found in the adherent cells. Indomethacin did not alter the suppressive effect. These studies demonstrate the induction of adherent suppressor cells in vitro and implicate the macrophage and CD4-positive T cells as the suppressor cells.

Clinical uses, collection, and banking of human milk.

The Mothers' Milk Bank (MMB) of San Jose, CA, has provided banked human milk to clinicians and researchers since its founding in 1974. This article briefly acknowledges the many and varied reasons for the use of human milk. Its primary focus however, is to review the protocol followed by the MMB for collection, banking, and distribution of human milk.

Inhibition of mouse natural killer cell activity by zinc.

The effect of zinc on mouse natural killer (NK) cell activity was evaluated. The inhibition of NK cell activity with zinc was dependent on the concentration of zinc added (range tested: 0-40 micrograms zinc/ml) and occurred at both effector-to-target ratios tested. Zinc-induced inhibition of NK activity was observed with the use of peritoneal or splenic effector cells on Toxoplasma gondii-augmented NK activity. Maximal inhibition of activity was noted when zinc was present for the entire assay period. Inhibition was present but less marked with pretreatment of effector cells with zinc. Pretreatment of target cells with zinc had no measurable effect on NK cytotoxicity. Effector-to-target cell binding as measured by single-cell assays was not significantly altered by zinc. These results indicate that zinc is a potent inhibitor of NK activity.

The bacterial content of breast milk after the early initiation of expression using a standard technique.

This study was initiated to evaluate the effect of early expression on the bacterial colony count of human milk. Significant bacterial contamination (greater than or equal to 10,000 colony-forming units/ml milk) was more common in 11 mothers who delayed the onset of expression of their milk compared with mothers who began to express their milk in the immediate postpartum period (n = 15) or who began to nurse their own full-term infants soon after delivery (n = 9). These data suggest that mothers who are separated from their prematurely born or sick infants should begin to express milk for their own infants as soon after birth as possible to provide milk with low bacterial contamination for frozen storage and later use.

Superior osteogenesis in transplanted allogeneic canine skull following chemical sterilization.

Sterilization of allogeneic bone increases the availability of this tissue for supplanting skeletal defects and effecting fusions. The optimal sterilant destroys micro-organisms, preserves the physical and chemical integrity of bone and possibly even reduces immunogenicity. Cortical bone of skull heals slowly and is variably resorbed. Of 36 dogs, spontaneous regeneration in 72 paired 20 mm defects was constant but always incomplete, and restored only about one third of the cross-sectional area of the defect at six months. The repair in defects replaced with canine allogeneic bony disc, sterilized with ethylene oxide (n = 9), gamma irradiation (n = 7), or methanol/chloroform/iodoacetic acid (n = 7) and then lyophilizedd, was compared with repair in defects filled with aseptically procured lyophilized only (n = 23) discs from the same donor. Criteria for evaluation of implants at six months included volume of defect filled, radiodensity, extent of fusion around circumference, revascularization, and remodeling. Bony discs sterilized with methanol/chloroform/iodoacetic acid remodeled at a superior rate (p less than 0.01). Radiation sterilization resulted in diminished density and inferentially reduced protection of the brain (p less than 0.025). Ethylene oxide, lyophilized implants, and implants lyophilized only produced comparable repair. Whereas an acceptable cranioplasty was achieved in 86% of methanol/chloroform/iodoacetic acid, lyophilize implants, all other alloimplants served an osteoconductive function with a successful repair occurring in 56% to 58%.

Ethylene oxide sterilization of bone, dura mater, and fascia lata for human transplantation.

The use of allogeneic human bone, dura, and fascia has achieved an enduring and accelerating role in the augmentation of spinal fusions and the repair of skeletal and dural defects. Primary sterilization of these nonviable cadaveric tissues magnifies the potential sources and ensures the microbiological sterility of the implant. Subsequent lyophilization facilitates preservation and distribution and reduces the immunogenicity of the graft. The evaluation of gaseous ethylene oxide (EO) as a sterilant was suggested by the delerious effects of alternative methods. Through a series of experiments, the following properties of EO sterilization were studied: (a) surface and interstitial sterilization; (b) the diffusion of EO into tissue, the formation of the reaction products ethylene chlorohydrin (EC) and ethylene glycol (EG), and the desorption of all three from tissues; (c) lyophilization and aeration in the removal of residues; and (d) minimization of residues through pretreatment. Gaseous EO is a very effective surface sterilant of wet bone, dura, and fascia and does not grossly alter these tissues. Its partial penetration through compact bone renders it less reliable for an interstitial antimicrobial effect, unless access to the interior is provided by serial openings. The toxicity of EO, EC, and EG mandates the desorption through lyophilization of these compounds (EC and EG are formed during sterilization with EO). Before sterilization, bone must be rid of marrow by vigorous irrigation with deionized water. The resultant reduction of the number of cells and of the available chloride decreases antigenicity and the formation of EC. Freeze-drying for more than 72 hours, in some cases augmented by prolonged aeration at room temperature, reduces EO, EC, and EG to acceptable levels. The accurate assay of residues in tissue requires acetone extraction for gas chromatography on rehydrated tissues because extraction of dry tissues gives falsely low results. Rigorous adherence to a protocol incorporating these findings justifies the acceptance of gaseous EO as a safe, relatively rapid, and inexpensive sterilant of bone and soft tissues.