Biomedical subjects
Publications and source records attributed to K Murray.
Protein B had a much higher affinity for human IgA than Jacalin, increasing the sensitivity and specificity of the measurement of total human IgA. Protein B, used as a capturing agent, greatly enhanced the measurement of antigen-specific IgA as compared to alpha chain-specific antibodies.
A monoclonal antibody SCH94.03, made in syngeneic mice by injection of spinal cord homogenate, promotes central nervous system remyelination when injected into SJL/J mice chronically infected with Theiler's virus. To elucidate the mechanism of antibody-mediated remyelination, SCH94.03 was investigated by immunocytochemistry, flow cytometry, immunoelectron microscopy, Western blotting, and immuno-thin layer chromatography. All cell types investigated in vitro showed strong cytoplasmic staining with a pattern resembling a cytoskeletal protein. In contrast, among the primary cultured cells studied, only oligodendrocytes showed strong surface reactivity. Other cell types, including astrocytes, microglia, Schwann cells, myoblasts, and T and B lymphocytes, were negative. Mouse and rat oligodendrocytes which showed strong surface reactivity exhibited a well-differentiated morphology, and approximately 50% expressed myelin basic protein. Since oligodendrocyte progenitors were negative for surface staining, the expression of the antigens recognized by this monoclonal antibody appears to be developmentally regulated, i.e., transiently expressed on younger, terminally differentiating oligodendrocytes. Among the cell lines studied, only two rat oligodendrocyte lineage cell lines showed surface reactivity with SCH 94.03. Western blotting of secondary isolated oligodendrocytes lysates revealed reactivity with multiple protein bands of 27, 32, 50, 100, and 106 kDa, whereas there was no reactivity to lipid antigens by immuno-thin layer chromatography. These results raise the possibility that SCH94.03 recognizes a novel oligodendrocyte-specific surface antigen, and may act directly on oligodendrocytes to promote remyelination.
IL-12 is a potent inducer of NK and cytolytic T cell activity, IFN-gamma production, and T cell proliferation, and is necessary for differentiation of naive T cells to the Th1 subset. We have previously shown that IL-12 promotes a primary Th1 response and suppresses a primary Th2 response in lymph nodes of mice primed with a model hapten-protein conjugate, 2,4,6-trinitrophenyl (TNP)-keyhole limpet hemocyanin (KLH). We have now extended these studies to determine the Th phenotype of the recall response following immunization with soluble Ag and IL-12. For these experiments, mice were primed with TNP-KLH with or without treatment with IL-12, allowed to progress beyond the primary immune response, and challenged by i.p. injection of TNP-KLH. The phenotype of the recall response was monitored by measuring ex vivo production of IFN-gamma and IL-4 in Ag-stimulated lymph node and spleen cell cultures. Titer and isotype of TNP-specific serum Abs were also evaluated. Mice primed with Ag+IL-12 developed a Th1 recall response, as detected by KLH-specific IFN-gamma production from cultured spleen cells and the presence of TNP-specific IgG2a Ab in serum. However, they also developed an Ag-specific Th2 recall response, as characterized by Ag-induced IL-4 production from spleen cells and the presence of high titers of anti-TNP IgG1 in the serum. Studies of the cytokine profile during the primary response revealed that IL-12 induced in spleen cells the capacity to express both IL-4 and IFN-gamma. CD4+ T cells are necessary for production of IL-4 in the spleens of IL-12-treated mice, and most likely account for the Th2 recall response detected in mice primed with Ag+IL-12. These results indicate that the Th1 phenotype induced by immunization with IL-12 and Ag is maintained so that a Th1 recall response is expressed upon subsequent challenge with Ag. However, immunization with IL-12 also supports the development of a Th2 recall response, indicating that the Th1-inducing effect of IL-12 in vivo is not accompanied by a long lasting suppression of Th2 development.
Fibroblast growth factors (FGFs) affect a broad spectrum of developmentally regulated cellular responses involved in the control of growth and differentiation. To identify specific FGF receptor forms involved in these responses, we have characterized FGF receptor transcript expression, and its modulation by FGF-2, as enriched populations of oligodendrocyte progenitors differentiate into mature oligodendrocytes. The data demonstrate that the levels of mRNA expression for FGF high-affinity receptors-1, -2, and -3 are differentially regulated during lineage progression: FGF receptor-1 expression increases with lineage progression, FGF receptor-2 is predominantly expressed by terminally differentiated oligodendrocytes, and FGF receptor-3 reaches a peak level of expression in late progenitors and then declines upon further differentiation; FGF receptor-4 expression was not detected in oligodendrocytes. Distinct patterns of alternatively spliced variants of FGF receptor-1 and -2 transcripts are expressed: the predominant FGF receptor-1 transcripts contain three Ig-like domains (FGF receptor-1 alpha), whereas the FGF receptor-2 transcripts contain two Ig-like domains (FGF receptor-2 beta 2) and this form is up-regulated as oligodendrocytes differentiate. In addition, the expression of these receptors is differentially regulated by the ligand, FGF-2: FGF receptor-1 mRNA expression is up-regulated in early progenitors, and FGF receptor-2 mRNA expression is down-regulated in mature oligodendrocytes. Finally, astrocytes express FGF receptor-1, -2, and -3 transcripts, but at different levels and with different exon utilization (FGF receptor-1 beta, FGF receptor-2 beta 1/beta 2) compared to oligodendrocytes. To our knowledge this is the first report that demonstrates that the mRNA expression of these three FGF receptor types is differentially regulated in primary cells as they differentiate along a lineage from progenitors to terminally differentiated cells. We propose that this pattern of expression provides a molecular basis for the developmentally varying response of cells to a common ligand. For example, according to this hypothesis, in response to FGF-2, FGF receptor-1 transduces signals that stimulate the prolonged proliferation and migration of early progenitors, FGF receptor-3 promotes the proliferation and arrest of differentiation of late progenitors, and FGF receptor-2 transduces signals for terminal differentiation, but not proliferation, in mature oligodendrocytes.
Differentiating cells undergo developmentally regulated changes in cell-cell and cell-matrix adhesion that control migration through microenvironments, proliferation, and differentiation. The diversity of the patterns of expression of heparan sulfate proteoglycans (HSPGs), coupled with their interactions with extracellular matrix, cell adhesion molecules, and growth factors, has emphasized their critical importance in the regulation of these events. Syndecans (1-4), glypican, and cerebroglycan are membrane-associated HSPGs that have been implicated in these events in various tissues and several tumor cell lines. We have examined the developmental expression and FGF-2-mediated regulation of these HSPGs during differentiation within a specific lineage of primary cells, oligodendrocytes (OL). Northern analyses of highly purified, developmentally synchronized populations of OL-lineage cells at three stages of differentiation (early and late progenitors and mature OLs) showed that the expression of individual forms of these syndecans and glypican are developmentally regulated. Specifically, the level of expression of syndecan-2 and -4 and glypican mRNAs increased as the cells differentiated from proliferative late progenitors to postmitotic mature cells. The expression of syndecan-1 and -3 had the inverse developmental pattern. Therefore, these two sets of molecules may have different roles in regulating the onset of terminal differentiation in OLs. The levels of mRNA expression were regulated by FGF-2: in late progenitors, FGF-2 induced a doubling of the mRNA levels of syndecan-2, -3, and -4, while those for syndecan-1 and glypican remained unaffected; in mature OLs, the levels of syndecan-1 mRNA were up-regulated, the levels of syndecan-2 and -4 and glypican were down-regulated. These results suggest that the individual syndecan molecules have distinct functions during the differentiation process and that multiple levels of regulation must exist, leading to a changing repertoire of these molecules during OL lineage progression and myelinogenesis.
Alternative donor bone marrow transplantation (BMT) to treat severe aplastic anaemia (SAA) in children and young adults has been complicated by high rates of graft rejection and severe graft-versus-host disease (GVHD). We hypothesized that increased immunosuppression combined with T-cell depletion of the marrow graft would enable successful use of unrelated donor BMT in this disease. Preconditioning consisted of cytosine arabinoside, cyclophosphamide, and total body irradiation (TBI). T-cell depletion was with the anti-CD3 antibody T10B9. GVHD prophylaxis consisted of cyclosporine A. 28 previously transfused patients were transplanted. Nine donor/recipient pairs were HLA matched. As of 1 January 1996, 15/28 (54%) patients are alive, transfusion independent and well with a range of follow-up of 13 months to 8 years (median 2.75 years). Fatalities include all three patients with nonengraftment and all three patients with grade III/IV GVHD. Other fatalities were due to infections or therapy-related toxicity. The incidence >or= grade II acute GVHD was 28%. These data show that in children with SAA who have failed immunosuppression, unrelated donor BMT offers a reasonable hope of long-term survival.
A case of isolated ureteral triplication is presented, the patient did not have any other urogenital anomaly. This presentation is rare and relevant literature is discussed.
PURPOSE: This study was a prospective phase I/II trial performed by the Radiation Therapy Oncology Group (RTOG) to test the tolerance and efficacy of preirradiation cyclophosphamide, doxorubicin, vincristine, and dexamethasone (CHOD) chemotherapy followed by large-volume, high-dose brain radiation therapy (RT) for patients with primary CNS lymphoma (PCNSL). PATIENTS AND METHODS: Fifty-four (52 assessable) human immunodeficiency virus (HIV)-negative patients with PCNSL were entered on study and received two (n = 20) or three (n = 32) cycles of CHOD (six patients with positive CSF cytology received intrathecal methotrexate in addition to CHOD). Whole-brain RT to 41.4 Gy and tumor boost to 18 Gy (total dose, 59.4 Gy) followed chemotherapy. RESULTS: As of July 1994, with a minimum potential follow-up time of 20 months, 12 of 52 assessable patients remain alive without evidence of progression. The median survival time for the entire group is 16.1 months, with a 2-year survival rate of 42%. By univariate analysis, patient age was found to be a significant prognostic factor with respect to survival (P = .005) in favor of age less than 60 years. Karnofsky performance status (KPS) was of borderline significance (P = .057). Survival for patients treated on RTOG 88-06 was compared with that of patients treated on RTOG 83-15, which tested RT alone. No difference in overall survival was found (P = .53). Grade 4 neutropenia developed in 29 of 51 patients during chemotherapy. There were two deaths during chemotherapy: one as a result of sepsis and one of a pulmonary embolus. The worst toxicity during RT was < or = grade 2 in 50 of 52 patients. CONCLUSION: Preirradiation CHOD chemotherapy does not significantly improve survival over RT alone for patients with PCNSL. Age remains a powerful prognostic factor independent of therapy and must be considered in testing alternative combined approaches.
PURPOSE: Definitive chemoradiation (CR) has replaced radical surgery as the preferred treatment of epidermoid carcinoma of the anal canal. To determine the importance of mitomycin (MMC) in the standard CR regimen and to assess the role of salvage CR in patients who have residual tumor following CR, a phase III randomized trial was undertaken by the Radiation Therapy Oncology Group (RTOG)/Eastern Cooperative Oncology Group (ECOG). PATIENTS AND METHODS: Between August 1988 and December 1991, 310 patients were randomized to receive either radiotherapy (RT) and fluorouracil (5-FU) or radiotherapy, 5-FU, and MMC. Of 291 assessable patients, 145 received 45 to 50.4 Gy of pelvic RT plus 5-FU at 1,000 mg/m2/d for 4 days, and 146 received RT, 5-FU, and MMC (10 mg/m2 per dose for two doses). Patients with residual tumor on posttreatment biopsy were treated with a salvage regimen that consisted of additional pelvic RT (9 Gy), 5-FU, and cisplatin (100 mg/m2). RESULTS: Posttreatment biopsies were positive in 15% of patients in the 5-FU arm versus 7.7% in the MMC arm (P = .135). At 4 years, colostomy rates were lower (9% v 22%; P = .002), colostomy-free survival higher (71% v 59%; P = .014), and disease-free survival higher (73% v 51%; P = .0003) in the MMC arm. A significant difference in overall survival has not been observed at 4 years. Toxicity was greater in the MMC arm (23% v 7% grade 4 and 5 toxicity; P < or = .001). Of 24 assessable patients who underwent salvage CR, 12 (50%) were rendered disease-free. CONCLUSION: Despite greater toxicity, the use of MMC in a definitive CR regimen for anal cancer is justified, particularly in patients with large primary tumors. Salvage CR should be attempted in patients with residual disease following definitive CR before resorting to radical surgery.
Patient with partial duplication of the distal penile urethra ending blindly in the periurethral tissue, without an external meatus is reported. Relevant literature and the classification of urethral duplication is discussed.
We examined inhibitory control as a quality of temperament that contributes to internalization. Children were assessed twice, at 26-41 months (N = 103) and at 43-56 months (N = 99), on repeated occasions, in multiple observational contexts and using parental reports. Comprehensive behavioral batteries incorporating multiple tasks were designed to measure inhibitory control at toddler and preschool age. They had good internal consistencies, corresponded with maternal ratings, and were developmentally sensitive. Individual children's performance was significantly correlated across both assessments, indicating stable individual differences. Girls surpassed boys at both ages. Children's internalization was observed while they were alone with prohibited objects, with a mundane chore, playing games that occasioned cheating, being induced to violate standards of conduct, and assessed using maternal reports. Inhibitory control was significantly associated with internalization, both contemporaneously and as a predictor in the longitudinal sense. The implications for considering children's temperament as a significant, yet often neglected contributor to developing internalization are discussed.
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OBJECTIVE: To describe the clinical and epidemiological features of an outbreak of a viral infection affecting humans and horses. SETTING: Stables in Hendra, a suburb of Brisbane. SUBJECTS: Affected horses and humans, and at-risk human contacts. RESULTS: A pregnant mare died two days after arrival from a paddock elsewhere in Brisbane. Eight to 11 days later, illness (depression, anorexia, fever, dyspnoea, ataxia, tachycardia, tachypnoea and nasal discharge) was reported among 17 other horses from the same or an adjoining stable. Fourteen horses died or were put down. Five and six days after the index mare's death, a stable-hand and then a horse-trainer, both of whom had had close contact with the sick mare's mucous secretions, developed influenza-like illnesses. The stable-hand recovered but the trainer developed pneumonitis, respiratory failure, renal failure and arterial thrombosis, and died from a cardiac arrest seven days after admission to hospital. A morbillivirus cultured from his kidney was identical to one isolated from the lungs of five affected horses. The two affected humans and eight other horses were seropositive for the infection, which was reproduced in healthy horses following challenge by spleen/lung homogenates from infected horses. There was no serological evidence of infection in 157 humans who had had contact with the stables or the sick horses or humans. CONCLUSIONS: A previously undescribed morbillivirus infected a probable 21 horses and two humans; one human and 14 horses died. That no further cases were detected among humans suggests that the virus was of low infectivity. The source of infection remains undetermined.
Studies of interactions between filamentous fusion phage particles and protein or nucleic acid molecules have gained increasing importance with recent successes of screening techniques based upon random phage display libraries (biopanning). Since a number of different phage are usually obtained by biopanning, it is useful to compare quantitatively the binding affinities of individual phage for the substrate used for selection. A procedure is described for determination of relative dissociation constants (KdRel) between filamentous phage carrying peptide fusions to the coat protein gpIII and substrates in solution. This novel method is based on the measurement of phage titres. Phage selected from a random fusion phage library for binding to a monoclonal antibody or a viral structural protein exhibited KdRel values in the nanomolar and micromolar ranges for their respective substrates, thus validating the method over a wide range of binding affinities.
Allogeneic bone marrow transplantation is the treatment of choice for many childhood leukemias. The donor of choice-an HLA matched sibling-is only available about 30% of the time. Unrelated donors are an alternative choice. In this report, we describe the results of unrelated donor bone marrow transplants (BMT) in 50 children with leukemia (25 acute lymphoblastic leukemia [ALL], 3 acute myeloid leukemia [AML], 3 juvenile chronic myelogenous leukemia [JCML], 10 chronic myeloid leukemia [CML]) or myelodysplastic syndrome (MDS; 9). The median age of the 31 male and 19 female patients was 9 years (range 2 to 18). Only 13 patients were serologically matched at HLA-A, B, DR, and DQ with their donors; 6 of these were reactive in mixed lymphocyte culture. The other 37 patients were mismatched for one (36 patients) or more (1 patient) HLA antigens. Pretransplant conditioning included cytosine arabinoside, cyclophosphamide, fractionated total body irradiation (TBI) (with lung, liver, and more recently, kidney shielding), and methylprednisolone. High-risk patients also received busulfan. Graft-versus-host disease (GVHD) prophylaxis consisted of T-cell depletion with IgM monoclonal antibody T10B9 plus complement and posttransplant cyclosporine-A. Forty-nine patients (98%) engrafted. Median times to greater than 500 polymorphonuclear leukocytes (PMN)/microL and greater than 25,000 platelets/microL were 18 and 20 days, respectively. Acute GVHD > or = grade II occurred in 16 patients (33%); 13 (81%) of these died. Chronic GVHD developed in 30 of 40 patients at risk, but was extensive in only 5. Event-free survival (EFS) for all patients was 44% +/- 7% (median follow-up was 49 months), and overall survival was 50 +/- 7%. Patients with low-risk disease (ALL or AML in first or second remission and CML in chronic phase) had a better EFS than children with high-risk disease (60% v 34%, P = .07). There was no significant difference in EFS between patients who were serologically matched with their donors (46%) and those who were partially mismatched (43%) (P = .97). These data compare favorably with published reports for children transplanted with HLA-matched sibling donors and should encourage earlier consideration of unrelated donor BMT in children with leukemia or myelodysplasia.
A morbillivirus has been isolated and added to an increasing list of emerging viral diseases. This virus caused an outbreak of fatal respiratory disease in horses and humans. Genetic analyses show it to be only distantly related to the classic morbilliviruses rinderpest, measles, and canine distemper. When seen by electron microscopy, viruses had 10- and 18-nanometer surface projections that gave them a "double-fringed" appearance. The virus induced syncytia that developed in the endothelium of blood vessels, particularly the lungs.
As an example for studies of contacts involved in complex biological systems, peptide ligands that bind to the core antigen of hepatitis B virus (HBcAg) have been selected from a random hexapeptide library displayed on filamentous phage. Affinity-purified phage bearing aa sequence LLGRMK, or some related sequences, bound full-length or truncated HBcAg but did not bind denatured HBcAg. The long (L), but not the short (S), hepatitis B virus envelope polypeptide, when synthesized in an in vitro system, bound firmly to HBcAg, indicating that interaction between HBcAg and the pre-S region of the L polypeptide is critical for virus morphogenesis. This interaction was inhibited by peptide ALLGRMKG, suggesting that this and related small molecules may inhibit viral assembly.
The epitope recognized by the monoclonal antibody MA18/7, specific for the PreS1-domain of the hepatitis B virus surface antigen, has been defined precisely by means of a library of fusion-phage carrying random hexapeptides on the tip of filamentous phage fd particles. Phage, isolated after only one round of affinity selection, displayed hexapeptides showing strong conservation of the PreS1 primary sequence in the region 19-23 with three noncontiguous residues, DP (20 and 21) and F (23) appearing in phage that bound the antibody. The importance of these core residues was supported by comparing the antibody binding of individual phage in solution, which provided relative dissociation constants for these interactions. Replacement of F (23) by Y was the only substitution observed in the three core residues, and resulted in somewhat weaker binding. Synthetic tetra- and hexapeptides containing these key residues inhibited the reaction between the phage and the antibody.