PubMed Health⌕ Search

Biomedical subjects

B H Johnson

Publications and source records attributed to B H Johnson.

At least 19 recordsLinked to original sources
PubMed2004

Identification of genes leading to glucocorticoid-induced leukemic cell death.

Glucocorticoidal steroids (GC) are capable of causing apoptotic death of many varieties of lymphoid cells; consequently, GC are used in therapy for many lymphoid malignancies. Gene transcription in the GC-treated cells is required for subsequent apoptosis, but only a few of the actual genes involved have been identified. We employed gene microarray analysis to find the network of genes involved in GC-evoked cell death, using three clones derived from the CEM lymphoid leukemia cell line. Clone C1-15 was resistant to GC-evoked apoptosis, although not necessarily to GC-induced gene transcription; the other two underwent apoptosis in the presence of GC. Clone C7-14 was subcloned from the apoptosis-sensitive parental C7 clone to establish karyotypic uniformity. The second sensitive clone, C1-6, was a spontaneous revertant from parental resistant clone C1. A period of > or = 24 h in the constant presence of receptor-occupying concentrations of synthetic GC dexamethasone (Dex) was necessary for apoptosis to begin. To identify the steps leading to this dramatic event, we identified the changes in gene expression in the 20-h period preceding the onset of overt apoptosis. Cells in the log phase of growth were treated with 10(-6) M Dex, and 2-20 h later, mRNA was prepared and analyzed using the Affymetrix HG_U95Av2 chip, containing probes for about 12,600 genes. Of these, approximately 6,000 were expressed above background. Comparisons of the basal and expressed genes in the three clones led to several conclusions: The Dex-sensitive clones shared the regulation of a limited set of genes. The apoptosis-resistant clone C1-15 showed Dex effects on a largely different set of genes. Promoter analysis of the regulated genes suggested that primary gene targets for GC often lack a classic GC response element.

Animals↗
PubMed2003

Evaluation of an extensive tuberculosis contact investigation in an urban community and jail.

SETTING: Urban community and jail. OBJECTIVES/DESIGN: Evaluate outcome and process of an extensive tuberculosis contact investigation, including completion of treatment of latent TB infection (TLTBI). RESULTS: Between April 2000 and September 2001, 18 epidemiologically-linked tuberculosis cases were identified; 15 were culture-confirmed, all with a matching 14-band DNA fingerprint pattern. The source case had cavitary pulmonary disease and had been incarcerated 4 months prior to diagnosis. Sixty-six of 67 (99%) community contacts and 221/344 (64%) jail contacts were evaluated. The presumed new infection rate was 56% for community contacts (11 cases, 25 tuberculin skin test [TST] positive) and 20% for jail contacts (6 cases, 32 TST converters). Screening results for 113 (33%) jail contacts were obtained in the jail TST registry upon rearrest. All identified cases completed treatment. Of 22 community contacts initiating TLTBI, 11 completed (44% of infected, 50% of initiators). Of 32 infected jail contacts, 12 initiated TLTBI (all who remained incarcerated), and 10 completed (31% of infected, 83% of initiators). None of 20 additional in-fected jail contacts, all of whose TST conversions were identified with re-arrest data, were subsequently located. Two additional related cases have been identified as of October 2003. CONCLUSIONS: Close health department/corrections collaboration facilitated this extensive contact investigation, which identified high Mycobacterium tuberculosis transmission rates and controlled the outbreak. Numerous contacts were identified and screened, but rates of treatment completion for infected contacts were low. Novel strategies are needed to maximize the number of infected contacts who are not only identified and evaluated, but completely treated.

Adolescent↗
PubMed2001

The pathway of leukemic cell death caused by glucocorticoid receptor fragment 465*.

The truncated glucocorticoid receptor mutant gene 465* codes for a protein that is interrupted by a frame-shift mutation in the second zinc finger of the natural DNA binding domain. Thus, 465* represents the natural amino acid sequence 1-465 followed by 21 novel amino acids starting at position 466. The entire ligand binding domain is missing. Prior studies have shown that transient transfection of the glucocorticoid-resistant leukemic T-cell clone ICR-27 with a plasmid expressing 465* rapidly reduces the number of viable cells. This response does not require activation by a steroid, and a hybrid protein consisting of green fluorescent protein fused to 465* is found primarily in the cytoplasm. In the present study, we present evidence that the decrease in cell number is due to a form of cell death that bears many of the classic characteristics of apoptosis. Expression of the 465* protein can be detected a few hours after electroporation and is followed by activation of caspase-3 as well as reduction of the mitochondrial inner transmembrane potential. The caspase-3 inhibitor ZVAD-fmk blocks 465*-dependent cell death when added acutely after electroporation, but fails to do so later. We conclude that the novel 465* gene causes cell death by apoptosis.

Apoptosis↗
PubMed2000

Structure-apoptotic potency evaluations of novel sterols using human leukemic cells.

Three oxidized analogs of cholesterol have been characterized for their ability to cause apoptotic cell death in CEM-C7-14 human leukemic cells. In addition to testing 15-ketocholestenol (K15), 15-ketocholestenol hydroxyethyl ether (CK15), and 7-ketocholesterol hydroxyethyl ether (CK7), an oxysterol of known apoptotic response, 25-hydroxycholesterol (25OHC), served as a standard for comparison. Growth studies based on dye exclusion by viable cells while using a sublethal concentration of oxysterols ranked their potency for cell kill as 25OHC > K15 > CK15 > CK7. Both the TUNEL assay (terminal deoxynucleotidyl transferase-mediated dUTP-X nick end labeling), which quantifies the amount of DNA nicks caused by a toxic agent, and the MTT assay, which measures cell metabolism and thus reflects cell viability, substantiated the same rank order. An ELISA assay for evaluating release of DNA fragments into the cytosol after treatment gave a similar potency order. The oncogene c-myc mRNA was suppressed by all three oxysterols, with 25OHC and K15 being the most potent suppressors. Hoechst and Annexin V staining documented that these oxysterols kill cells by an apoptotic pathway as evidenced by condensation of nuclear chromatin and plasma membrane inversion, respectively. From these in vitro studies, we believe that 25OHC, K15, and possibly CK15 have the potential to be chemotherapeutic agents.

Apoptosis↗
PubMed2000

Transfer student and gender issues within the Department of Animal Science at Texas A&M University.

Graduates with a B.S. or graduate degree from the Department of Animal Science (1986 to 1996) of Texas A&M University were surveyed by mail to gather opinions on the curriculum and satisfaction with their degrees. The survey contained five Likert questions (1 = excellent, 4 = poor) regarding satisfaction with major, educational results, personal growth, professional development, and skills important for employment. Multiple-choice questions (n = 9) addressed education, background, job status, and gender. Surveys (n = 2,192) were sent to students, and 470 responded. The undergraduate respondents (n = 396) comprised 71.2% (n = 282) nontransfer students and 28.8% (n = 114) transfer students. Transfer students reported a mean grade point ratio (GPR) of 3.05 +/- 0.43 at Texas A&M University in comparison to a nontransfer student mean GPR of 3.03 +/- 0.45 for hours completed at Texas A&M University (P = 0.83). The results for time from graduation to full-time employment for transfer students were similar to those of nontransfer students (P = 0.95). Differences in current annual salary between transfer students and nontransfer students were apparent (P < 0.0001). Transfer students and nontransfer students reported similarly that courses within the department were valuable (P = 0.95) and not redundant (P = 0.08). The two groups similarly rated contributions to critical thinking (P = 0.59) and speaking (P = 0.38). Undergraduate respondents (n = 392) were 48.9% (n = 192) male and 51.1% (n = 200) female. Males and females reported a mean GPR of 3.02 +/- 0.46 and 3.07 +/- 0.43, respectively (P = 0.35). Differences in job search time (P < 0.0001), starting salary (P = 0.0004), and current salary (P < 0.0001) were apparent between genders; females were paid less and searched longer before finding their first job. Males and females responded similarly that courses were valuable (P = 0.16) and were taught effectively (P = 0.37) and that teaching assistants made positive contributions (P = 0.43). Females felt a lesser contribution to critical thinking ability, leadership ability, and technical expertise. Results suggest the continuance of current educational practices, but changes to specific issues should be initiated.

Adult↗
PubMed1998

Resistance of human leukemic CEM-C1 cells is overcome by synergism between glucocorticoid and protein kinase A pathways: correlation with c-Myc suppression.

Glucocorticoids (GCs) induce apoptosis in lymphoid cells that contain functional GC receptors (GRs). However, GC resistance often is seen in cells with demonstrable GRs; one such line is CEM-C1. We have tested the hypothesis that positive interactions between GC and cyclic AMP (cAMP) regulate GC actions in CEM clones. Treatment of both GC-resistant CEM-C1 [resistant to 1 microM dexamethasone (Dex)] and the sensitive sister clone, CEM-C7 (approximately 65% cell death with 20 nM Dex, approximately 99% death with 1 microM Dex), with a < or = 20 microM concentration of the protein kinase A activator, forskolin, had no significant effect on cell viability. Cotreatment with Dex and forskolin resulted in a strong synergistic death response, with only approximately 10% CEM-C1 cells surviving treatment with 1 microM Dex and 20 microM forskolin. This death was blocked by the GR antagonist RU 38486. However, the extent of apoptosis did not correlate with the amount of GR protein or binding activity in either C7 or C1 cells. As reported previously, Dex-evoked cell death was associated with suppression of c-Myc in C7 cells. In CEM-C1 cells, Dex alone did not affect c-Myc; however, Dex plus forskolin suppressed c-Myc levels. To evaluate mechanisms of Dex-forskolin synergism, fresh subclones of CEM-C7 (clone 14) and CEM-C1 (clone 15) were isolated, to ensure purity of phenotype. In these, forskolin (with or without Dex) caused a similar increase in cAMP (approximately 300-fold) and phospho-cAMP-responsive element binding protein (approximately 4-5-fold) levels, whereas total cAMP-responsive element binding protein expression was not affected. GR transcription function, as tested from a GR-responsive 330-bp mouse mammary tumor virus promoter-luciferase reporter construct, was induced 8- and 4-fold by 1 microM Dex treatment of CEM-C7-14 and CEM-C1-15 cells, respectively. Forskolin (10 microM) significantly potentiated Dex response in CEM-C1-15 cells (13.5-fold) but had only a modest effect (1.5-fold) in CEM-C7-14 cells. These studies suggest that sensitization of CEM-C1 cells by cross-talk between GR and protein kinase A pathways may occur via cooperative effects on GR-mediated gene transcription.

Apoptosis↗
PubMed1997

Characteristics of 25-hydroxycholesterol-induced apoptosis in the human leukemic cell line CEM.

Cholesterol and related compounds can give rise to oxygenated sterol molecules (oxysterols) which are potent regulators of lymphoid cell growth. Oxysterols added exogenously cause cell death of several lines of cultured cells, and on the basis of limited criteria, it has been suggested that this death is apoptosis. In the present study, we show definitive evidence that 25-hydroxycholesterol (25OHC) kills cells of the clone CEM-C7 by apoptosis and establish the temporal sequence of related cellular and biochemical events. Cell shrinkage was evident as early as 12 h, while cell death was not evident until after 24 h. It mounted rapidly after that, and by 72 h, virtually all cells were dead. Electron microscopic analysis shows that by 24 h after treatment and before the onset of cell death, early ultrastructural features typical of apoptosis were present. DNA breaks were detected by TUNEL assay prior to the onset of cell death. Two types of specific DNA pieces often associated with apoptosis were found as increasing numbers of cells died. DNA fragments of 300 and 50 kbp were not appreciable until 42 h, and internucleosomal cleavage was observed by 48 h after oxysterol addition. None of these effects were seen in an oxysterol-resistant CEM subclone, establishing the specificity for apoptosis of the biochemical and morphological events. z-VAD.FMK, a peptide inhibitor of ICE-related proteases delayed but did not prevent the apoptosis of CEM-C7 cells induced by 25OHC. The addition of mevalonate partially protected CEM-C7 cells from apoptosis but did not restore cell growth.

Amino Acid Chloromethyl Ketones↗
PubMed1997

Glucocorticoid/oxysterol-induced DNA lysis in human leukemic cells.

Both glucocorticoids and oxysterols, steroids with quite different known transduction pathways, cause the death of lymphoid cells. Dual TUNEL/propidium iodide assays on sensitive human leukemic CEM-C7 clones treated with either steroid were clearly positive by 48 h, consistent with apoptosis. Both steroids evoked two distinctive types of DNA lysis: cleavage into large fragments of several different sizes and the classic "ladders", multiples of approximately 200 base pairs. Conventional gel electrophoresis showed that a small proportion of total DNA had undergone laddering 36-48 h after treatment with glucocorticoid or 24 h after oxysterol exposure. On field inversion gel electrophoresis of cellular DNA both steroids caused an increase in an array of large DNA fragments <50 kb in size. A 50 kb fragment appeared 36 h after treatment with either steroid, but only oxysterol treatment caused a significant increase in a 300 kb fragment. Oxysterol treatment did not result in DNA fragmentation in the resistant M10R5 subclone, which retained sensitivity to glucocorticoids. We conclude that glucocorticoids and oxysterols kill these cells with similar, but not identical, patterns of DNA lysis which occur just before or concomitant with the onset of cell death.

Cell Division↗
PubMed1996

Clinical value of radiologists' interpretations of perioperative radiographs of orthopedic patients.

The content and accuracy of radiographic interpretations by radiologists was assessed to determine the reports' ability to provide sufficient information necessary to make clinical treatment decisions. A retrospective review was performed of 371 radiographic studies (211 consecutive patients) and their reports which had been generated by three Board-certified radiologists. Data were collected regarding fracture assessment and description as well as the description and assessment of orthopedic implants. These descriptions were categorized by their ability to be used clinically (precise) or not (general). Fracture descriptions were considered complete for 85% of reports, while their assessment of alignment and displacement (necessary to determine fracture care) was complete on only 9%. Orthopedic implants were described precisely on 12% of reports with 7% of the descriptions in error. The effect and position of orthopedic implants were described precisely for only 27% and 25% of cases, respectively, while implant stability was assessed precisely in only 4% of cases. For 61% of preoperative studies, the radiologist's report was not available until after the surgical procedure had already been performed. For all variables considered, an average of 3% of descriptions contained an error. Radiologists' reports of radiographs of these patients undergoing orthopedic procedures did not contain sufficient descriptive information to be used clinically, were not promptly available, and contained an error for 3% of variables studied. The attending orthopedic surgeon has traditionally interpreted such radiographs and should continue to do so to provide patients with more immediate and complete clinical evaluation and management.

Fractures, Bone↗
PubMed1996

Transfected glucocorticoid receptor and certain GR fragments evoke cell death in malignant lymphoid, not myeloid cell lines.

Previously we have shown that glucocorticoid sensitivity could be restored to a clone of glucocorticoid-resistant leukemic T cells by transfecting them with an expression vector for the glucocorticoid receptor. Furthermore, transfection with plasmids expressing fragments of the receptor containing the DNA-binding domain resulted in constitutive loss of cells. In this paper, we report the results of transfecting both types of constructs into lines of glucocorticoid-resistant human leukemic cells of T cell, B cell, and myeloid origin. In all the lymphoid lines tested, transfection of the holoreceptor gene resulted in appearance of steroid-dependent cell death. In the same lines, transfection of glucocorticoid receptor fragments expressing amino acids 1-465* (465 residues of the normal sequence plus a novel 21 amino acid C-terminus) or expressing only 398-465* caused cell death without the addition of steroids. The amount of cell loss following transfection of these constitutively lethal fragments was in the same range as that following transfection of the holo glucocorticoid receptor plus administration of glucocorticoid. However, the cell loss due to the constitutively active fragments occurred more rapidly. Neither of the myeloid lines tested were sensitive to any of the transfected constructs, with or without added steroid.

Apoptosis↗
PubMed1995

Glucocorticoid antagonist RU 486 reverses agonist-induced apoptosis and c-myc repression in human leukemic CEM-C7 cells.

A synthetic glucocorticoid dexamethasone (DEX) inhibits proliferation and induces apoptosis of clone CEM-C7 cells, but not in clone CEM-C1 cells, even though they contain glucocorticoid receptors (GR). We previously showed that suppression of c-myc is a critical step in glucocorticoid-induced cell lysis of C7 cells. It is not reduced in C1 cells. In this study we review the basis for this conclusion and present evidence that the glucocorticoid antagonist RU 486 rescues DEX-treated C7 cells from cell death. An increase in DEX-repressed c-myc mRNA levels precedes the recovery of cell growth. A threshold level of Myc expression appears to be required to maintain growth and viability of C7 cells. Although C1 cells are highly resistant to lysis by glucocorticoids, addition of forskolin, an inducer of protein kinase A, synergizes to evoke complete apoptosis. This synergistic effect is prevented by RU 486, indicating direct involvement of the GR.

Apoptosis↗
PubMed1994

Effects of consuming endophyte-infected tall fescue on growth, reproduction and lactation in mice selected for high fecundity.

Effects of a diet containing endophyte-infected tall fescue seed (83% infected) were investigated using 2 lines of mice, one line selected for fecundity (L(+)) and the other a randomly selected control line (K). Treatments included a commercial stock diet (C), 50% stock plus 50% non-infected tall fescue seed (N), and 50% stock plus 50% infected tall fescue seed (I). The experiment was conducted using mice on respective treatments in 2 phases (successive generations), with 15 to 23 mated females per line and diet subgroups. Mated females of Phase 1 were assigned at random within line to experimental diets which were fed during gestation and through 21 d of lactation. Litters were standardized to 10 pups 1 d after birth. Stock diets were fed to all groups from Day 21 to weaning on Day 28. Weaned male and female pups were allotted to previous diets. Mated females in Phase 2 were managed as in Phase 1 through weaning at 28 d. Diets of males did not affect reproduction and data were pooled within female diets. Selected (L(+)) dams gave birth to more live pups than K dams (P<0.05) during both phases (+3.4 and +2.8 +/- 0.4 pups, respectively). Diet but not line affected littering rate of mated females in Phase 1 (71.3%, I; < 87.1%, C or 93.0%, N; P<0.05) and Phase 2 (82.1%, I < 93.8%, N or 97.1%, C; P<0.05). Diet had no effect on fecundity during Phase 1 but females on I diet had reduced (P<0.05) litter size by 1.9 and 3.2 +/- 0.5 pups compared with the females on N and C diets, respectively, in Phase 2. Feed consumption and weights of dams during lactation generally ranked C>N>I. Growth of pups during both phases also ranked C>N>I. Vaginal opening at 28 d differed by line (71.4%, K < 89.3%, L(+), P<0.05) and diet (56.8%, I < 92.0%, C or 92.2%, N, P<0.05). These results suggest both acute and chronic effects of consumption of endophyte-infected diets. Absence of line-by-diet interactions demonstrates that adverse effects were unrelated to genetic differences between lines.

Journal Article↗
PubMed1994

Oxysterol sensitive and resistant lymphoid cells: correlation with regulation of cellular nucleic acid binding protein mRNA.

Oxygenated derivatives of cholesterol inhibit cholesterol synthesis, prevent lymphoid cell growth, and evoke cell death. We have employed a novel selection method to isolate M10 cells, a line of oxysterol-resistant cells, from the sensitive clone CEM C7. Concentrations of the potent sterol 25-hydroxycholesterol that occupy the oxysterol binding protein cause cell death in CEM C7, but not in M10 cells. Both cell lines have similar amounts of the oxysterol binding protein with similar affinities for oxysterol. However, in neither line are the levels of oxysterol binding protein mRNA affected by 1 microM 25-hydroxycholesterol. Furthermore, both cells express the cellular nucleic acid binding protein (CNBP), a 7 zinc finger, DNA-binding protein of unknown function, regulated by oxysterols. The levels of CNBP mRNA are significantly reduced by 25-hydroxycholesterol in the sensitive CEM C7 cells, in which the dose response and time course are consistent with occupancy of the oxysterol binding protein by oxysterol and with subsequent cell kill. However, in the resistant M10 cells, CNBP mRNA levels are unaffected by these concentrations of the 25-hydroxycholesterol. Our results suggest a role for CNBP in oxysterol-induced regulation of cell viability and growth.

Cell Death↗
PubMed1994

Recombinant proteins can be isolated from E. coli cells by repeated cycles of freezing and thawing.

Repeated cycles of freezing and thawing are sufficient to separate highly expressed recombinant proteins away from the cellular milieu of E. coli. Freezing and thawing liberates recombinant proteins from the bacterial cytoplasm, but does not release the bulk of endogenous E. coli proteins. Furthermore, protein secretion is not required. Fractionation of overexpressed proteins by freeze/thaw treatment does not depend on the identity of the recombinant protein and has been observed for thirty-five different recombinant proteins expressed in E. coli. These include proteins originally found in plant, animal or microbial sources, as well as several proteins designed de novo. Freezing and thawing typically yields approximately 50% of the recombinant protein in relatively pure form. Thus the freeze/thaw treatment can be utilized as a general method for the isolation of recombinant proteins from E. coli.

Chemical Fractionation↗
PubMed1993

Autonomic characteristics of nonclinical panic and blood phobia.

Autonomic characteristics of nonclinical panic and blood phobia were compared using spectral analysis of the electrocardiogram (EKG), as well as more conventional cardiovascular measures. The cardiovascular responses of 11 subjects who reported recent occurrence of frequent severe panic attacks, and 10 subjects who reported intense somatic reactions to the sight of blood (including episodes of syncope) were recorded during a variety of laboratory tasks (quiet rest, reaction time/shock avoidance, face immersion, and combined reaction time/face immersion). Results suggest distinct autonomic patterns in the groups. Panickers showed (a) higher heart rate and reduced heart-rate variability (b) aberrant associations among cardiovascular measures, and (c) dominant sympathetic control of heart rate coupled with diminished vagal tone. Blood phobics generally displayed an opposite pattern. The relevance of these findings to the etiology of panic and blood phobia, as well as to biological models of anxiety disorders in general, is discussed.

Adult↗
PubMed1993

Oxysterol-induced cell death in human leukemic T-cells correlates with oxysterol binding protein occupancy and is independent of glucocorticoid-induced apoptosis.

In eukaryotic cells oxysterols inhibit cholesterol biosynthesis and cell growth. A potent oxysterol, 25-hydroxycholesterol, was used to investigate the biological effects of oxysterols on three clonal lines of either glucocorticoid-sensitive or -resistant CEM cells, human leukemic T-lymphocytes. In addition, the glucocorticoid sensitivity of an oxysterol-resistant CEM cell line was tested. Oxysterols blocked growth and caused the lysis of cells regardless of their glucocorticoid response. All cells studied herein possessed an oxysterol binding protein with high affinity for 25-hydroxycholesterol. For all clones grown in serum-free medium, the half-maximal cytolytic concentration of 25-hydroxycholesterol (20-40 nM) correlated with its affinity (Kd = approximately 31 nM) for this oxysterol binding protein. Both cholesterol and mevalonate reversed 25-hydroxycholesterol cytotoxicity; 3-6 microM cholesterol or 0.1 mM mevalonate decreased 60 nM 25-hydroxycholesterol cytotoxicity by 50%. This cholesterol or mevalonate reversal appeared possible even after several days of 60 nM oxysterol treatment. The protective effect of cholesterol could be overcome by increasing 25-hydroxycholesterol concentrations. Cholesterol and mevalonate did not prevent glucocorticoid-mediated lymphocytolysis. Furthermore, the oxysterol-resistant line was sensitive to dexamethasone lysis. These data support the hypothesis that oxysterols and glucocorticoids act independently to block the growth of human leukemic lymphoblasts.

Apoptosis↗
PubMed1993

The effects of background visual roll stimulation on postural and manual control and self-motion perception.

The effects of background visual roll stimulation on postural control, manual control, and self-motion perception were investigated in this study. In the main experiment, 8 subjects were exposed to wide field-of-view background scenes that were tilted and static, continuously rotating, or sinusoidally rotating at frequencies between 0.03 and 0.50 Hz, as well as a baseline condition. The subjects performed either a postural control task (maintain an upright stance) or a manual control task (keep an unstable central display horizontally level). Root-mean square (RMS) error in both the postural and manual control tasks was low in the static tilt condition and extremely high in response to continuous rotation. Although the phases of the postural and manual responses were highly similar, the power and RMS error generated by the sinusoidal visual background stimulation peaked at a lower frequency in the postural task. Vection ratings recorded at the end of the postural and manual trials somewhat paralleled the frequency tuning differences between tasks, which a subsequent experiment showed to be the result of the differential motion of the central display rather than the differential positioning of the subject. In general, these results show that the dynamic characteristics of visual orientation systems vary according to the specific motor and/or perceptual system investigated.

Adult↗