Biomedical subjects
Publications and source records attributed to W H Knospe.
Hematologic emergencies require thorough but prompt evaluation of the patient, availability of relatively routine laboratory tests, and immediate decisions. This article discusses the emergencies associated with red blood cell disorders, white blood cell disorders, hemostatic disorders, and transfusion reactions.
Cellulose ester membranes (CEM) were coated with stromal cells from murine bone or bone marrow irradiated in vitro with 1000, 2000, or 4000 rad and then implanted i.p. in CAF1 mice for periods of six and 12 months. CEM coated with stromal cells from bone showed excellent regeneration of bone and hematopoiesis after 1000 rad in vitro irradiation. After 2000 rad, hematopoietic and bone regeneration was reduced by about 50%, and after 4000 rad it was completely absent in CEM coated with stromal cells from bone. CEM coated with stromal cells from bone marrow showed no regeneration of hematopoiesis or bone after 1000, 2000, and 4000 rad in vitro irradiation and residence i.p. for six and 12 months. These results indicate that regeneration of the hematopoietic microenvironment is dependent upon living stromal cells. A difference in radiation sensitivity is demonstrated between stromal cells from bone and from bone marrow.
The ultrastructure of developing osseous and hematopoietic tissue and supporting stroma was examined within intraperitoneally implanted, marrow-coated cellulose ester membranes (CEMs). During initial periods of implantation (two weeks), coated CEMs were shallowly infiltrated and surface-lined with mainly two stromal cell types--primitive mesenchymal cells and large, pleomorphic, multinucleated monocytoid-like stromal cells--and, in addition, endothelial cells in early stages of vessel development. Selective proliferation, maturation, and orientation of these cell types along the CEM's surface (one month), resulted in the formation of primitive osseous and hematopoietic tissue sites. Osseous sites developed as undifferentiated mesenchymal cells transformed into well-differentiated secretory cells residing in an electron-dense, pre-mineralized extracellular matrix, that upon extended implantation (3-6 months) formed bone. Hematopoietic sites developed as mesenchymal cells, in intimate contact with the monocytoid-like stromal cells, extended elongated branches into the medullary cavity, and enveloped newly formed surface-associated vascular structures. With subsequent maturation of vascular sinuses and supporting adventitial stroma (3-6 months), the sites were colonized with either uni-, bi-, or trilineal hematopoietic elements. These observations provide evidence that common, marrow-derived precursor stromal cells, i.e., mesenchymal and multinucleated monocytoid-like cells, cooperate in the development of both osseous and hemic tissue sites.
A 45-year-old white male, bisexual, with a 2-year history of acquired immunodeficiency syndrome (AIDS) prodrome, developed a Burkitt cell-like acute lymphoblastic leukemia (ALL). Marker studies of marrow blasts show an unusual and possibly unique pattern, in that an unequivocal monoclonal B cell leukemia, having K-IgM with HLA-DR and B cell subset antigen (BA-1) expression, was superimposed with a mature suppressor T cell marker profile (pan-T, mature T, and suppressor/cytotoxic T antigens). The leukemic blasts were totally negative for terminal deoxynucleotidyl transferase (TdT), human T cell leukemia-lymphoma virus (HTLV) p19 antigen, and other immunoglobulin isotypes. Chromosome analysis of marrow cells disclosed that 70% of the cells had 47,XY, + 12,t(8;14)(q24;q32) chromosome complement, and 30% of the cells had a 47,XY, + 12,dup1q + (q22-31),t(8;14)(q24;q32). The consistent finding of the specific chromosome rearrangement (8/14 translocation) in all abnormal cells suggests that the cells were derived from a common precursor. With regard to the partial T cell marker expression, the significance of these markers in B cell leukemia is unclear, as is their relation to the additional chromosome abnormalities that apparently developed in the process of clonal evolution.
Implants of bone and tooth matrix powder were placed subcutaneously (s.c.) on intraperitoneal (i.p.) Mitex or Polyvic membranes. Implants were removed for histology after 1-24 weeks. Macrophages, fibroblasts, and vascular sinusoids infiltrated around bone and tooth matrix particles after one week. In the s.c. tooth matrix implants, a few sites of cartilage formation and ossification developed at two weeks, and by three weeks granulocytopoiesis and megakaryocytopoiesis developed adjacent to new bone; erythropoiesis was not observed. In s.c. bone matrix implants and in the i.p. artificial membranes coated with bone or tooth matrix powder, ossification or hematopoiesis was not observed. Small numbers of CFU-s, CFU-nm, BFU-e, and CFU-e appeared 10-20 days after s.c. implantation of tooth matrix; none were detected in s.c. bone matrix implants.
Cellulose ester membranes (CEM) were implanted intraperitoneally into S1/S1d and S1+/ S1+ mice. These CEM rapidly became coated with peritoneal cells capable of supporting primarily granulocytic colony development after seven days. S1/S1d-coated CEM showed a diminished capacity to support colony development compared with S1+/ S1+ CEM, perhaps reflecting the defect in the hematopoietic microenvironment of these mice. Marrow cells from S1+/S1+ and S1/S1d mice generated similar numbers of colonies on S1+/S1+ CEM. When CEM were transferred from a primary to a secondary host there was a tendency to remodel the CEM toward the characteristics of the secondary host. Peritoneal cells coating CEM from S1/S1d mice had less phagocytosis of yeast particles than peritoneal cells from S1+/S1+ mice. The cell coat on the membranes from S1/S1d mice was fewer cell layers in thickness than those on membranes coated in S1+/ S1+ mice.
Conventional treatment of polycythemia vera (PV) with radioactive phosphorus or alkylating agents is associated with a significant excess of acute leukemia and cancer of the gastrointestinal tract and skin. There is thus a need for a nonmutagenic agent in the treatment of this disorder. Hydroxyurea (HU) was administered to 118 patients with a loading dose of 30 mg/kg/day for 1 week, which was then reduced to 15 mg/kg/day. Initial control of the elevated hematocrit and platelet count was achieved within 12 weeks in over 80% of patients. Long-term disease control was defined and the accumulative 1-year failure-free survival was 73% in the previously untreated patients and 59% in those patients previously treated with other myelosuppressive modalities. The HU was well tolerated and cytopenia, which generally occurred within the first 8 weeks of therapy, was transient and of little clinical significance. However, it is recommended because of this toxicity that HU be administered initially at a dose of 15-20 mg/kg/day. Three patients developed acute leukemia; two were untreated and one had had myelosuppressive therapy. Hydroxyurea is an effective agent in the treatment of PV, but continued assessment of its mutagenic potential is necessary.
Artificial membranes composed of mixed cellulose nitrate and acetate esters (MCEM), polytetrafluoroethylene (Mitex), cellulose acetate (Celotate), polyvinylchloride (Polyvic), polycarbonate (Nuclepore), or linear polyethylene with pore sizes ranging from 0.45 mu to 70 mu were coated on their inner surfaces with a paste of bone marrow, diaphyseal bone, or cells proliferating in vitro on the inner surfaces of curetted femurs obtained from syngeneic mice. Squares of these membranes were folded into thirds, stapled to form an open-ended, tube-like structure, and implanted i.p. into mice. At intervals of 6-12 months, the membranes were removed and studied histologically. Control membranes not coated with any substance were also studied. Trilineal hematopoiesis, bone formation, sinusoidal structures, fat cells, and hemosiderin-laden macrophages were found with all three coatings in the tubes formed from the MCEM or Mitex membranes. The Celotate and Polyvic membranes coated with bone cells and the Polyvic membranes coated with marrow also developed trilineal hematopoiesis and new bone tissue. The remaining membranes essentially had only fibrous tissue, except for rare foci of hematopoiesis which developed on an occasional 30 mu (but not 70 mu) pore size linear polyethylene membrane. The results of these studies suggest that hematopoiesis and bone cell formation on these artificial membranes is closely linked, and that there may be some interaction between membrane constituents and the cells destined to form the hematopoietic support layer in the fostering of hematopoiesis. Whether this interaction is stimulatory or inhibitory was not defined by these studies.
Immunologically mediated aplastic anemia (AA) in mice was induced by the i.v. injection of 10(7) lymph node cells (LNC) from H-2k identical but Mls mismatched CBA/J donor mice into previously irradiated (600 rad total body gamma) C3H/HeJ mice. Cyclosporin A (CsA), 25 mg/kg, was administered subcutaneously from day -1 to day 30. Control mice included C3H/HeJ mice which received 600 rad alone, C3H/HeJ mice which received 600 rad plus CsA as above, and C3H/HeJ mice which received 600 rad total body irradiation followed by 10(7) LNC from CBA/J donors. CsA failed to prevent lethal AA. These results suggest that the pathogenetic mechanisms operating in immunologically mediated AA differ from the mechanisms operating in rodents transplanted with allogeneically mismatched marrow or spleen cells which develop graft-versus-host disease. The results are consistent with a non-T cell-dependent mechanism causing the AA.
Histochemical procedures were used to elucidate differences in the distribution of glycosaminoglycans (GAGs) and glycoproteins (GPs) in tubes made of cellulose-ester membrane (CEM) in six weeks and 12 months following intraperitoneal (i.p.) implantation. After six weeks, CEM tubes coated on their inner surfaces with bone marrow (BM), bone homogenate (BH), or regenerating medullary mesenchyme (M) contained a highly vascularized loose connective tissue and scattered hematopoietic elements. Numerous mononuclear cells were seen at the stromal-CEM interface and infiltrating the CEM. At the site of infiltration, there were accumulations of GPs and nonsulfated acidic GAGs. The remainder of the CEM contained limited amounts of sulfated and nonsulfated GAGs. By 12 months, both coated and uncoated CEMs were saturated with sulfated and nonsulfated acidic GAGs. However, only coated CEM tubes contained trilineal hematopoiesis surrounded by a shell of bone that was incorporated into the structure of the CEM. Exterior to the bone, the CEM was infiltrated with GPs, some of which were acidic and nonsulfated, possibly sialoglycoproteins. No bone or glycoproteins infiltrated the uncoated CEMs. The stroma supporting trilineal hematopoiesis contained GPs and nonsulfated acidic GAGs, but not sulfated acidic GAGs. However, both sulfated and nonsulfated GAGs were found in the fibrous connective tissue filling the lumens of uncoated CEMs. While CEM tubes accumulate peritoneal GAGs that may be conducive for the retention, differentiation, and proliferation of osteogenic and other stromal cells provided by coatings of BM, BH, or M, it is these cells that produce the bone and glycoproteins that appear to contribute to a microenvironment conducive to trilineal hematopoiesis. However, the relative roles of these substances in this process remain to be elucidated.
The search for cures for the various forms of cancer has long occupied the forefront of medical research. In leukemia, bone marrow transplantation is effecting cure in more and more types of leukemia and in different stages of the disease. This comprehensive review provides the reader with the background and rationale for transplantation, a description of the procedure itself and the potential complications, and a look at the state of the art.
Cellulose ester membranes (CEM) of pore size 0.45 mu were enriched on their inner surfaces with spleen or bone marrow or bone or regenerating primitive endosteal tissue folded into a tubular configuration and implanted i.p. Although CEM with enrichment with bone or marrow or spleen supported trilineal hematopoiesis 3 to 6 months after i.p. implantation, CEM at 8 and 9 months were progressively fibrotic and involuted with minimal hematopoietic support capacity. CEM of pore size 3 mu were enriched with bone marrow or bone or regenerating endosteal tissue. These CEM after 9 and 12 months supported excellent trilineal hematopoiesis and were always associated with osteogenesis. Regenerating primitive endosteal tissue is an equally effective source of stroma to regenerate the hematopoietic microenvironment. Unenriched CEM became progressively more fibrotic after 6 months.
Immunologically mediated aplastic anemia (AA) was experimentally induced in mice by injecting 10(7) lymph node cells (LNC) from donor mice of one inbred strain to another H-2k identical but Mls mismatched strain previously given 600 rad total body gamma irradiation (TBI). AA developed after 2 weeks to 6 months in selected strain combinations used and usually 60 to 90% of the mice died. Clinical signs of graft-versus-host disease did not occur and splenic atrophy rather than splenomegaly was the rule. Histologically these mice had a lesion of the hematopoietic microenvironment characterized by sinusoidal injury and stromal necrosis. Others have demonstrated injury to hematopoietic stem cells. C3H/He LNC induced AA whereas C3H/HeJ LNC failed to induce AA. The C3H/HeJ strain carries a macrophage defect and these results suggest that a macrophage-like cell may be a mediator of immunological injury in this experimental model. Although all strain combinations evaluated were H-2k identical and Mls mismatched, certain Mls combinations resulted in AA and identical Mls mismatched but different strains did not. Both strong (Mlsd) and weak (Mlsc) stimulating LNC induce AA but simple Mls differences do not explain the AA as similar Mls combinations but different strain combinations fail to induce AA.
Mouse bone matrix powder was implanted subcutaneously or put into folded tubular cellulose ester membranes (CEM) and implanted i.p. Calcification and new bone formation did not develop. Sinusoidal vascularization of the matrix developed by 3 to 4 wk. Stromal cells such as fat cells and fibroblasts were present by 4 wk. Hematopoiesis was absent except for rare foci of granulopoiesis on some of the CEM at 6 wk. Rat bone matrix implanted subcutaneously into mice and mouse bone matrix implanted subcutaneously into rats failed to induce new bone formation or hematopoiesis.
Thirty-one patients with essential thrombocythemia were randomized to receive either melphalan or radioactive phosphorus as myelosuppressive therapy. Twenty-seven patients were evaluable for response. Of 13 patients treated with melphalan, 11 had a complete response (platelet count less than 450,000/mm3) at 3 and 6 months. This response rate was significantly better than the response to radioactive phosphorus. The response rates were similar at 12 months. No significant toxicity was observed with either regimen.
Uracil mustard was used for the treatment of thrombocytosis in 14 patients, eight who had polycythemia vera and six who had essential thrombocytosis. Intermittent treatment with 1 to 2 mg/day for 14 days was used for most patients, and continuous treatment was used for three patients. The nadir of the platelet count occurred during a mean period of five to seven weeks, and the mean duration between courses was seven months. The drug selectively depressed the platelet count, with a minimal effect on leukocytes and erythrocytes. Uracil mustard is an effective drug for the control of thrombocytosis.
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Two patients with thrombotic thrombocytopenic purpura (TTP) were treated with plasmapheresis procedures in which the replacement fluid included normal plasma. In both cases, the platelet count rose, hemolysis decreased, and neurologic symptoms improved promptly after this therapy. Unfortunately, both patients eventually died, one of complications of end-organ damage and the other of complications of maintenance therapy with prednisone. Plasmapheresis may be an effective treatment for TTP, either by removing a toxic factor or supplying a deficient factor.