Analysis of 135 autopsy eyes for ocular involvement in leukemia.

To explain the marked variation in the reported incidence of how often leukemic cells infiltrate the eye in fatal cases of leukemia, we tested the hypothesis that ocular leukemic infiltration is related to the peripheral leukocyte count during the final hours of life. We reviewed tissue sections, as well as autopsy and clinical records, from 135 patients who had fatal leukemia and had their eyes examined after death at Duke University Medical Center. Infiltrates of leukemic cells were found in the eyes of 42 of 135 patients (31.1%), with the choroid being the most frequently involved site. We detected a significant positive correlation between the ocular leukemic infiltration and an agonal leukocyte count as well as the severity of systemic disease. Differences in the agonal circulating leukocyte count may partly explain variations in the incidence of leukemic infiltrates in different postmortem studies.

Interactions between spinach ferredoxin-nitrite reductase and its substrates. Evidence for the specificity of ferredoxin.

Reduced ferredoxin can serve as electron donor in the 6-electron reduction of nitrite to ammonia catalyzed by spinach nitrite reductase. We have examined interactions between nitrite reductase and its substrates, ferredoxin and nitrite, with emphasis upon protein-protein interactions between ferredoxin and nitrite reductase. Ferredoxin, of the proteins tested, is the most effective in retarding low ionic strength inactivation of nitrite reductase. The interaction appears to be electrostatic, and the apparent Kd, calculated from the concentration dependence of ferredoxin protection, is about 1 microM in 2 mM Tris. Chemical modification of carboxyl residues of ferredoxin resulting in a change of charge reduces its reactivity with both ferredoxin:NADP+ oxidoreductase and nitrite reductase, indicating the importance of charge-charge interactions. Cross-linking studies provided no evidence for a ternary complex containing the oxidoreductase and nitrite reductase but indicated that the two enzymes will compete for ferredoxin, possibly using the same site (or overlapping sites) on the ferredoxin. A complex containing ferredoxin:NADP+ oxidoreductase, ferredoxin, and cytochrome c was detected, indicating that ferredoxin has different binding sites for cytochrome c and ferredoxin:NADP+ oxidoreductase. Active cross-linked complexes of ferredoxin and nitrite reductase were obtained and were less sensitive to low ionic strength inactivation than free reductase and had decreased ferredoxin-supported nitrite reductase activity. The evidence presented of protein-protein interactions between ferredoxin and nitrite reductase indicates that ferredoxin is indeed the specific physiological electron donor to the reductase.

Intercellular relationships in the synthesis of macromolecules by organ cultures of corneas.

Sepharose CL-4B chromatography of guanidine hydrochloride and aqueous extracts of 3H-glucosamine labeled intact corneal tissue reveals four peaks representing proteoglycans and glycoproteins. To evaluate the universality of the 4th peak, hereafter designated as Sepharose CL-4B (IV), its presence was investigated in rabbit, bovine, cat, rhesus monkey, and human corneal preparations. Following incubation in isotopically labeled medium, corneas were extracted with aqueous and/or 4M guanidine hydrochloride and subjected to Sepharose CL-4B chromatography. Sepharose CL-4B (IV) was detected in all species studied; 3H-glucosamine and 14C-amino acids, but not 35SO4, were incorporated into this peak which eluted in the range consistent with an apparent molecular weight of approximately 30,000 D. To determine which layers were involved in the synthesis of Sepharose CL-4B (IV) the layers of the rabbit cornea were incubated separately (stroma scraped of endothelium and/or epithelium, epithelium only, endothelium only). A distinct Sepharose CL-4B (IV) peak was not identified in the chromatographs obtained from organ cultures of corneal epithelium, endothelium, or from corneal stroma scraped of epithelium and/or endothelium. This decrease in Sepharose CL-4B (IV) synthesis occurred even if the scraped cornea was not allowed to expand in volume by compressing it beneath a membrane porous to the incubation medium. Thus, Sepharose CL-4B (IV) synthesis was enhanced significantly by the stroma being in conjunction with other corneal cells as they exist in vivo.