George F. Tidmarsh

Bone marrow stromal cell lines with lymphopoietic activity express high levels of a pre-B neoplasia-associated molecule

Bone marrow stromal cell lines have been isolated that directly support B lymphopoiesis in vitro. Single B-lineage precursors proliferate and differentiate on certain of these stromal cell lines to establish long-term B-lineage cultures. These lymphopoietic stromal cells produce novel soluble factors that support proliferation of in vitro established pre-B cell populations. Lymphoid populations established on lymphopoietic stromal cell lines lack surface Ig-bearing cells, but give rise to surface Ig+ cells when transferred to mixed bone marrow feeder layers. Several stromal lines expressed a B-lineage neoplasia marker detected by the monoclonal antibody MAb6C3. Remarkably, only the 6C3Aghi stromal lines supported long-term proliferation of B-lineage cells. We propose that the 6C3 antigen-bearing molecule may play a role in stromal cell-dependent, pre-B cell proliferation, as well as in neoplastic proliferation of pre-B leukemias.

Angiotensin II for the Treatment of Vasodilatory Shock

Background Vasodilatory shock that does not respond to high‐dose vasopressors is associated with high mortality. We investigated the effectiveness of angiotensin II for the treatment of patients with this condition. Methods We randomly assigned patients with vasodilatory shock who were receiving more than 0.2 μg of norepinephrine per kilogram of body weight per minute or the equivalent dose of another vasopressor to receive infusions of either angiotensin II or placebo. The primary end point was a response with respect to mean arterial pressure at hour 3 after the start of infusion, with response defined as an increase from baseline of at least 10 mm Hg or an increase to at least 75 mm Hg, without an increase in the dose of background vasopressors. Results A total of 344 patients were assigned to one of the two regimens; 321 received a study intervention (163 received angiotensin II, and 158 received placebo) and were included in the analysis. The primary end point was reached by more patients in the angiotensin II group (114 of 163 patients, 69.9%) than in the placebo group (37 of 158 patients, 23.4%) (odds ratio, 7.95; 95% confidence interval [CI], 4.76 to 13.3; P<0.001). At 48 hours, the mean improvement in the cardiovascular Sequential Organ Failure Assessment (SOFA) score (scores range from 0 to 4, with higher scores indicating more severe dysfunction) was greater in the angiotensin II group than in the placebo group (‐1.75 vs. ‐1.28, P=0.01). Serious adverse events were reported in 60.7% of the patients in the angiotensin II group and in 67.1% in the placebo group. Death by day 28 occurred in 75 of 163 patients (46%) in the angiotensin II group and in 85 of 158 patients (54%) in the placebo group (hazard ratio, 0.78; 95% CI, 0.57 to 1.07; P=0.12). Conclusions Angiotensin II effectively increased blood pressure in patients with vasodilatory shock that did not respond to high doses of conventional vasopressors. (Funded by La Jolla Pharmaceutical Company; ATHOS‐3 ClinicalTrials.gov number, NCT02338843.)

Normal thymic cortical epithelial cells developmentally regulate the expression of a B-lineage transformation-associated antigen.

Nonlymphoid, stromal cells in the mouse thymus are believed to be important in T cell maturation and have been proposed to play a central role in the acquisition of major histocompatibility complex (MHC) restriction and self-tolerance by maturing thymocytes. Both cortical and medullary epithelial cells in the thymus express high levels of class II (A) major histocompatibility antigens (MHC Ags). We show here that a specific subset of these A− epithelial cells express a transformation-associated antigen (6C3Ag) found previously on the surfaces of Abelson murine leukemia virus-transformed pre-B cells and on those bone marrow-derived stromal cell clones which support normal and preneoplastic pre-B cell proliferation. Among solid lymphoid organs, only the thymus contains 6C3Ag1 cells and within the thymus, this antigen is found exclusively on A− epithelial cells in cortical regions. It is striking that the expression of the 6C3Ag on thymic epithelium is developmentally regulated, suggesting a role for this lymphostromal antigen in the maturation of the thymic microenvironment.

Identification of a novel bone marrow-derived B-cell progenitor population that coexpresses B220 and Thy-1 and is highly enriched for Abelson leukemia virus targets.

A novel stage in early B-lymphocyte differentiation has been identified in normal mouse bone marrow cells. Earlier work had demonstrated that bone marrow cells characterized by low levels of Thy-1 and lack of a panel of lineage markers (Thy-1lo Lin- cells) were highly enriched for pluripotent hematopoietic stem cells. In this paper, we present evidence that another bone marrow population, which expressed low levels of Thy-1 and coexpressed B220, a B-lineage-specific form of the leukocyte common antigen, contained early and potent precursors for B lymphocytes upon in vivo transfer to irradiated hosts. These Thy-1lo B220+ cells, comprising 1 to 2% of bone marrow cells, were enriched for large cells in the mitotic cycle; the population lacked significant pluripotent hematopoietic stem cell activity and myeloid-erythroid progenitors. Most strikingly, Thy-1lo B220+ cells represented a highly enriched population of bone marrow cells that could be targets of Abelson murine leukemia virus transformation. We propose that Thy-1lo B220+ bone marrow cells represent the earliest stage of committed lymphocyte progenitors, intermediate in differentiation between Thy-1lo Lin- pluripotent stem cells and, in the B lineage, Thy-1- B220+ pre-B cells.

Rapid release of Iron from Ferritin by Siderophores

The ability of the microbial siderophores deferriferrichrome, deferriferrichrome A, and enterobactin to remove iron from ferritin has been investigated. In contrast to previously published data with other chelators, all three siderophores rapidly released iron from the mammalian storage protein. Enterobactin was found most efficient at removing ferritin-bound iron. Using this siderophore, the mechanism by which ferritin sequesters iron was studied. The relative iron saturation level of ferritin influenced the rate of chelation by the microbial siderophores.

The in vitro translation product of the murine λ 5 gene contains a functional signal peptide

Abstract We have isolated and sequenced a novel λ 1 constant region related cDNA clone which might represent an allelic variant of the recently described λ 5 gene. This λ 5 transcript is present in pre-B cell lines and bone marrow cells, but not in B cell lines, plasma cell lines or in spleen cells. In vitro translation studies show that the translation product contains a signal peptide of approx. 30 amino acids at its N -terminus.

Acquisition of iron from transferrin bySalmonella paratyphi B

The ability ofSalmonella paratyphi B to obtain iron from human, bovine, and rabbit transferrin was studied. All three transferrins restored the growth potential for otherwise ironstarvedS paratyphi B producing enterobactin. No exogenous siderophore was needed for growth to occur. A non-enterobactin-producing derivative ofS. paratyphi B was unable to acquire iron from transferrin, and transferrin did not restore the growth potential of the organism. Sequestering the transferrin fromS. paratyphi B in dialysis tubing did not affect the ability of the transferrin to supply iron to the organism.

Pharmacodynamic evaluation of intragastric pH and implications for famotidine dosing in the prophylaxis of non-steroidal anti-inflammatory drug induced gastropathy—a proof of concept analysis

Abstract Objective: Famotidine given at a dose of 80 mg/day is effective in preventing NSAID-induced gastropathy. The aim of this proof of concept study was to compare twice a day (BID) vs 3-times a day (TID) administration of this total dose of famotidine on intragastric pH in healthy volunteers. Research design and methods: Two analyses were undertaken: (1) a 13 subject controlled cross-over 24-h intragastric pH evaluation of the BID and TID administration of 80 mg/day of famotidine, as well as measures for drug accumulation over 5 days (EudraCT, number 2006-002930-39); and (2) a pharmacokinetic (PK)/pharmacodynamic (PD) model which predicted steady-state famotidine plasma concentrations and pH of the two regimens. Results: For the cross-over study, gastric pH was above 3.5 for a mean of 20 min longer for TID dosing compared to BID dosing on Day 1. On Day 5, the mean time above this threshold was higher with the BID regimen by ∼25 min. For pH 4, subjects’ gastric pH was above this pH value for a mean of 25 min longer for TID dosing compared to BID dosing on Day 1. For Day 5, the pH was above 4 for ∼45 min longer with the TID regimen as compared with the BID regimen. The mean 24-h gastric pH values when taken in the upright position trended higher for the TID dosing period compared to the BID regimen on Day 1. The steady-state simulation model indicated that, following TID dosing, intragastric pH will be above 3 for 24 h vs 16 h for the BID regimen. There was no evidence for plasma accumulation of famotidine with TID dosing as compared to BID dosing from either analysis. Conclusion: The data indicate that overall more time is spent above the acidic threshold pH values when 80 mg/day of famotidine is administered TID vs BID. Key limitations included small study size with a short duration and lack of a baseline examination, but was compensated for by the cross-over and PK/PD modeling design. Although most of the comparisons in this proof of concept study were not statistically significant these results have important implications for future research on gastric acid lowering agents used for the prevention of NSAID-induced gastropathy.

Angiotensin converting enzyme defects in shock: implications for future therapy

Background Patients who develop vasodilatory shock, particularly when caused by an inflammatory condition like sepsis or pancreatitis, have evidence of significant endothelial injury as manifested by coagulation disorders and increased capillary permeability [1, 2]. Since angiotensin converting enzyme (ACE) activity is primarily endothelium membrane-bound [3], patients with vasodilatory shock may develop an ACE defect [4, 5]. The pulmonary and renal capillary beds hold the majority of endothelium-bound ACE and patients with acute respiratory distress syndrome (ARDS) have increasing ACE insufficiency with increased severity of lung injury [5, 6]. Moreover, previous studies have demonstrated that endotoxemia causes a decrease in ACE function [7, 8], and, finally, ACE function has been shown to be important in sepsis outcomes [5, 9, 10]. Based on these findings, investigators from the first ATHOS trial have suggested that endothelial dysfunction in vasodilatory shock may cause a significant ACE defect that results in angiotensin II (ANG-2) insufficiency [4].

Normal and Neoplastic Early Lymphocyte Maturation

T and B lymphocytes apparently share a common precursor with other hematopoietic cells in the bone marrow. Using surface phenotype-defined bone marrow cells and clonogenic assays for pre-T and pre-B cells we have defined several stages of early lymphocyte maturation by bone marrow cells. We have examined the maturation of lymphocytes in Whitlock-Witte cultures, and have defined 2 stages of early lymphocyte maturation prior to the expression of the B cell form of the lymphocyte common antig. i (B220). The earliest of these two stages residues in a population of Thy-110 cells which make up only 0.1% of bone marrow cells, yet contain all known precursors for the various hematolymphoid lineages, including pluripotential stem cells. These cells undergo early maturation on certain cloned stromal elements (e.g. AC 6.21) from the Whitlock-Witte cultures, reaching the stage of expression of the B220 antigen, but prior to the expression of surface immunoglobulin. Transfer of the cells from cloned stromal cells to miced stromal elements in the Whitlock-Witte culture system gives rise to surface Ig+ B cells in a short time, indicating at least 2 stromal elements necessary for the proliferation and differentiation of 2 major stages of B lymphocyte maturation. Abelson leukemia virus transformed pre-B cells require stromal elements in the early phases of leukemogenesis for their growth, and the cloned stromal line (AC 6.21) responsible for the earliest B lymphocyte maturation from the Thy-110 cells is sufficient for this task. Later, at about the time the Abelson pre-B transformants become feeder-layer independent, ley begin to express a tumor -associated B 1ineage antigen which we call gp1606C3.