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Biomedical subjects

A K Smith

Publications and source records attributed to A K Smith.

At least 19 recordsLinked to original sources
PubMed2007

Glucocorticoid receptor polymorphisms and haplotypes associated with chronic fatigue syndrome.

Chronic fatigue syndrome (CFS) is a significant public health problem of unknown etiology, the pathophysiology has not been elucidated, and there are no characteristic physical signs or laboratory abnormalities. Some studies have indicated an association of CFS with deregulation of immune functions and hypothalamic-pituitary-adrenal (HPA) axis activity. In this study, we examined the association of sequence variations in the glucocorticoid receptor gene (NR3C1) with CFS because NR3C1 is a major effector of the HPA axis. There were 137 study participants (40 with CFS, 55 with insufficient symptoms or fatigue, termed as ISF, and 42 non-fatigued controls) who were clinically evaluated and identified from the general population of Wichita, KS. Nine single nucleotide polymorphisms (SNPs) in NR3C1 were tested for association of polymorphisms and haplotypes with CFS. We observed an association of multiple SNPs with chronic fatigue compared to non-fatigued (NF) subjects (P < 0.05) and found similar associations with quantitative assessments of functional impairment (by the SF-36), with fatigue (by the Multidimensional Fatigue Inventory) and with symptoms (assessed by the Centers for Disease Control Symptom Inventory). Subjects homozygous for the major allele of all associated SNPs were at increased risk for CFS with odds ratios ranging from 2.61 (CI 1.05-6.45) to 3.00 (CI 1.12-8.05). Five SNPs, covering a region of approximately 80 kb, demonstrated high linkage disequilibrium (LD) in CFS, but LD gradually declined in ISF to NF subjects. Furthermore, haplotype analysis of the region in LD identified two associated haplotypes with opposite alleles: one protective and the other conferring risk of CFS. These results demonstrate NR3C1 as a potential mediator of chronic fatigue, and implicate variations in the 5' region of NR3C1 as a possible mechanism through which the alterations in HPA axis regulation and behavioural characteristics of CFS may manifest.

5' Flanking Region↗
PubMed2004

Freeze-substitution and low-temperature embedding of dairy products for transmission electron microscopy.

Dairy products are comprised largely of fat, air and water, which makes it difficult to preserve their ultrastructure for electron microscopy. Keeping the samples frozen throughout fixation and embedding protects the structure and distribution of the components of emulsions and foams. Therefore, dairy products were freeze-substituted and embedded at low temperature (-20 degrees C) to prepare them for transmission electron microscopy. Whipped cream, ice cream mix and dairy/non-dairy mixed systems were frozen by plunging in propane, at its boiling point (-187 degrees C). Ice cream, because it is already frozen, was fractured into 1-mm3 pieces in liquid nitrogen and then added to frozen fixative (-196 degrees C). Fixative solution consisted of glutaraldehyde, osmium tetroxide and uranyl acetate dissolved in either methanol or acetone. When material was to be stained after sectioning the fixative was limited to glutaraldehyde in methanol. The temperature was increased step-wise from -80 to -20 degrees C. Solvent was replaced with resin; the polar resin Lowicryl HM4, the non-polar resin Lowicryl HM20, LR White and LR Gold were tested. Samples were embedded and polymerized at -20 degrees C using ultraviolet light to cross-link the resin. Methanol proved to be the most effective solvent for substituting the ice; the hydrophobic resin Lowicryl HM20 was the most effective resin for retaining fat structure following osmium fixation.

Journal Article↗
PubMed2003

Flow cytometric characterization of perfused human bone marrow cultures: identification of the major cell lineages and correlation with the CFU-GM assay.

BACKGROUND: Prolific cultures of human bone marrow mononuclear cells (BM MNCs) were recently developed that include a full spectrum of hematopoietic and accessory cells, with the presence of autofluorescent cells indicating adequate cell expansion. However, phenotypic and functional clonogenic characterizations of the autofluorescent cells and the various other subpopulations present in these cultures have not been carried out. METHODS: Cells from a continuously perfused bioreactor inoculated with BM MNCs and cultured for 12 days in serum-containing medium with PIXY321, erythropoietin, and with or without FLT3-L were evaluated by using flow cytometry. RESULTS: Two antibodies, CD71 and CD13, allowed the separation of the autofluorescent cells into two distinct populations. The CD71+CD13++ autofluorescent population contained the colony-forming unit (CFU) fibroblast, and the CD71++CD13++ autofluorescent population contained macrophage/dendritic like cells. The CFU-granulocyte/macrophage (CFU-GM) could not be thoroughly evaluated with CD71 and CD13. However, the number of CD13+/++Lin- cells correlated with the number of CFU-GM (r = 0.83), with approximately 1 CFU-GM for every 30 CD13+/++Lin- cells. CONCLUSIONS: The data showed that CD71 and CD13 antibodies separate the autofluorescent cells into two populations but do not separate hematopoietic cells into specific phenotypic populations. The data also showed that the number of CD13+/++Lin- cells correlated with the number of CFU-GM. These data present the initial step toward detailed phenotypic analysis of ex vivo expanded human BM MNC cultures.

Antigens, CD↗
PubMed2002

Pregnancy rates for grade 2 embryos following administration of synthetic GnRH at the time of transfer in embryo-recipient cattle.

To succeed with pregnancy a bovine embryo must overcome the luteolytic mechanism and achieve recognition of pregnancy. It is understood that well developed embryos are more successful in achieving recognition of pregnancy than poorly developed ones. Attempts have been made to assist this recognition of pregnancy by utilising a number of hormonal supplements with varying levels of success. A study was undertaken to test the hypothesis that supplementation with synthetic GnRH at the time of transfer of Grade 2 embryos will enhance pregnancy rates in recipients receiving this category of embryo. Pairs of fresh and frozen Grade 2 embryos (n = 38) from 34 donor animals were allocated to the trial. Thirty eight pairs of recipients were used and one of each pair was randomly assigned to receive treatment on the day of embryo transfer (Day 7) with 5 ml of gonadorelin, containing a synthetic gonadotrophin releasing hormone, 0.1 mg/ml. Pregnancy diagnosis was carried out from 42 days post-transfer by either palpation per rectum or ultrasound scanning. Treatment, embryo processing, side of transfer, parity of recipient, breed of recipient and breed of donor dam showed no statistically significant effect on pregnancy rate. The overall pregnancy rate in this study was within commercially accepted limits for Grade 2 embryos at 38.2%. The pregnancy rates were 34.2 and 42.1% for the GnRH-treated and control groups, respectively and were not significantly different at P < 0.05. The failure of this treatment to improve pregnancy rates could be due to its effect being transitory therefore allowing subsequent pregnancy loss. The timing of the treatment post-transfer, treatment dose and potency of the GnRH analogue may also play a role in this. Further study is required to determine the hormonal or follicular status of prospective candidates for treatment before applying this as a whole herd regime.

Animals↗
PubMed2002

Transforming growth factor beta 1 dysregulation in a human oral carcinoma tumour progression model.

A human oral tumour progression model was established that consists of normal epithelial cells and three cell lines representing stages from dysplastic to metastatic cells. To investigate the impact of exogenous transforming growth factor-beta 1 on this model system, we analysed the responsiveness of those cells to transforming growth factor-beta 1 and explored the potential mechanism underlying the transforming growth factor-beta 1 activity. We found that the growth of all cell types, regardless of their stage of tumour progression, is inhibited by transforming growth factor-beta 1, although to different degrees. Transforming growth factor-beta 1 induced the expression of cyclin-dependent kinase inhibitors p15(INK4B), p21WAF1/(CIP1) and p27(KIP1). In contrast, transforming growth factor-beta 1 was found to stimulate the invasive potential of one cell type that represents the most advanced stage of tumour phenotype, suggesting that the impact of transforming growth factor-beta 1 on functional features of tumour cells other than cellular proliferation may play a significant role in the process of oral tumour progression.

Active Transport, Cell Nucleus↗
PubMed2002

Plasma membrane disruption in orthodontic tooth movement in rats.

Sublethal plasma membrane disruption (PMD) is an established mechanism for signaling in several cell types, including endothelial cells and skeletal muscle. We used a rat model of orthodontic tooth movement to test the hypothesis that periodontal ligament (PDL) cells communicate stretch to changes in bone cell activity in part via PMD. To produce PMD, we used a 50-g load from a spring activated in the buccal direction against the maxillary first molars for 5 min. Uptake of endogenous serum albumin was used as a PMD marker. Immunohistochemistry demonstrates albumin in PDL cells surrounding moved first molar tips. Image analysis shows significantly more albumin in cells of the buccal side (tension) of the moved teeth compared with those of the lingual, distal, and mesial sides, and those of the unmoved control. Albumin localization within cells of the PDL, after only 5 min of mechanical loading, suggests that PMD could promote uptake or release of signaling molecules.

Analysis of Variance↗
PubMed2001

Effects of overexpression of gamma-Glutamyl hydrolase on methotrexate metabolism and resistance.

Intracellular metabolism of methotrexate (MTX) to MTX-polyglutamates (MTXPG) is one determinant of cytotoxicity. Steady-state accumulation of MTXPG seems to depend on the activity of two enzymes: folylpolyglutamate synthetase (FPGS), which adds glutamate residues, and gamma-glutamyl hydrolase (GGH), which removes them. Overexpression of GGH would be expected to decrease intracellular MTXPG, thereby increasing efflux of MTX and decreasing cytotoxicity. Increased expression of GGH has been shown to be associated with resistance to MTX in human sarcoma cell lines and a rat hepatoma cell line. To clarify the specific role of GGH in determining MTX sensitivity, we investigated the phenotype produced by forced GGH overexpression in two cell types. Furthermore, because MTX and folic acid share metabolic pathways, we measured the effects of GGH overexpression on folic acid metabolism. The full-length cDNA for GGH, subcloned into a constitutive expression vector, was transfected into a human fibrosarcoma (HT-1080) and a human breast carcinoma (MCF-7) cell line. Compared with the clones containing an empty vector, the GGH-overexpressing cells express 15- to 30-fold more GGH mRNA, more GGH protein, and 15- to 90-fold more GGH enzyme activity. GGH overexpression altered MTX accumulation and metabolism to long-chain polyglutamates. In contrast to expectations, however, GGH overexpression did not confer resistance to short MTX exposures in either cell line. Changes in MTX metabolism were found to be balanced by alterations in accumulation and metabolism of folic acid. The ratio of MTX:folate accumulation may be a better predictor of MTX cytotoxicity than the accumulation of either alone. We conclude that, at least for these two cell lines, GGH overexpression alone is insufficient to produce clinical resistance to MTX.

Antimetabolites, Antineoplastic↗
PubMed2001

Radiographic findings and Gs-alpha bioactivity studies and mutation screening in acrodysostosis indicate a different etiology from pseudohypoparathyroidism.

Acrodysostosis is an uncommon skeletal dysplasia associated with nasal hypoplasia, midface deficiency, severe brachydactyly, and varying degrees of hearing loss and mental retardation. Previous publications have suggested that it may be difficult to distinguish acrodystostosis from pseudohypoparathyroidism on clinical grounds, but acrodysostosis does appear to have distinct clinical and radiologic findings. Spinal stenosis is an underappreciated risk in acrodysostosis, despite the reported loss of normal caudal widening of the lumbar interpediculate distance on AP spine radiographs in the original report of this disorder by Robinow et al., with confirmation of these radiographic findings by Butler et al. We report two sporadic cases of acrodysostosis, one of which required decompressive laminectomy for symptomatic spinal stenosis, and review 11 cases of acrodysostosis from 9 families that were submitted to the International Skeletal Dysplasia Registry. The objective of this report is to determine the frequency and severity of spinal stenosis in patients with acrodysostosis and to summarize the clinical and radiographic findings of acrodysostosis in an effort to distinguish acrodysostosis clearly from pseudohypoparathyroidism. The pattern of brachydactyly differs between these two conditions, and varying degrees of spinal stenosis are characteristic of acrodysostosis. Both our index patients with acrodysostosis had normal bioactivity of the alpha subunit of the Gs protein, therefore indicating that acrodysostosis has a different pathogenesis from pseudohypoparathyroidism. Furthermore, single-strand confirmational polymorphism (SSCP) analysis failed to demonstrate any confirmational alterations in the coding exons of the Gs alpha gene. These radiographic and laboratory findings substantiate that acrodysostosis is clinically different from pseudohypoparathyroidism and that it is necessary to follow patients with acrodysostosis for signs of spinal stenosis.

Dysostoses↗
PubMed2001

A phase II trial evaluating the safety and effectiveness of the AastromReplicell system for augmentation of low-dose blood stem cell transplantation.

To reduce the number of apheresis procedures and maintain the usual rate of hematopoietic recovery in patients treated with high-dose chemotherapy, we studied the effect of adding a small volume of ex vivo expanded bone marrow to low doses of CD34(+) blood stem cells. Thirty-four patients with breast cancer received G-CSF (10 microg/kg/day) priming followed by a limited volume (50-100 ml) bone marrow aspiration and standard 10-liter aphereses. Marrow was expanded ex vivo using the AastromReplicell system and infused along with low doses of blood-derived CD34(+) cells, collected in one apheresis. Thirty-one evaluable patients received a median CD34(+) blood stem cell dose of 0.7 x 10(6)/kg (range, 0.2-2.5) and 4.7 x 10(7) nucleated cells/kg (range, 1.98-8.7) of ex vivo expanded marrow. All patients recovered with normal blood counts and engrafted 500 neutrophils/microl and 20 000 platelets/microl in a median of 10 and 13 days, respectively. Multivariate analysis revealed that, in addition to CD34(+) lineage negative cell quantity, the quantity of stromal progenitors contained in the ex vivo expanded product correlated with engraftment outcome (r = 0.551, P = 0.004). Our results indicate that ex vivo expanded bone marrow is capable of facilitating engraftment when combined with low doses of mobilized blood derived CD34(+) cells.

Adult↗
PubMed2001

Dicarbonyldi-mu-chloro-cis,cis-eta4-1,5-cyclooctadienedirhodium(I).

The title compound, dicarbonyl-1kappa(2)C-di-mu-chloro-1:2kappa(4)Cl-[cis,cis-2(eta(4))-1,5-cyclooctadiene]dirhodium(I), [Rh(2)Cl(2)(C(8)H(12))(CO)(2)], consists of a dichloro-bridged dimer of rhodium, with a non-bonded Rh.Rh distance of 3.284 (2) A. One Rh atom is coordinated to two carbonyl ligands, while the other Rh atom is coordinated to the cyclooctadiene moiety.

Journal Article↗
PubMed2001

Connexin-43 expression in oral-derived human osteoblasts after transforming growth factor-beta and prostaglandin E2 exposure.

Dental implant placement stimulates a response in the supporting tissue; the response involves bone remodeling and release of wound-healing factors, including cytokines. Important factors such as transforming growth factor-beta (TGF-beta), which promotes matrix synthesis, and prostaglandin E2 (PGE2), a mediator of inflammation, have the potential to alter the communication between bone cells and interfere with implant site healing. Cells responsible for the formation of bone are interconnected to form a multicellular network. Cell-to-cell communication in this network occurs in part via gap junctions. In bone cells, the predominant gap junction protein is connexin-43. TGF-beta is a growth modulator produced by osteoblasts and released from the matrix in response to resorption and may influence the progression of periodontal disease. TGF-beta also promotes the synthesis of extracellular matrix proteins such as collagen, fibronectin, and adhesion molecules. PGE2 is a mediator of inflammation produced in response to periodontal pathogens. PGE2 levels in the gingival sulcular fluid have been correlated with attachment loss and bone resorption. The relationship between these factors and connexin-43 is unclear. Oral-derived (alveolar) bone was used because the phenotype of bone can differ between species and between different sites in the body. For our studies, explants of human osteoblasts were cultured on eight well plates and characterized by their expression of osteocalcin, osteonectin, alkaline phosphatase, type 1 collagen, and connexin-43. Cells were grown to near confluence on 12 well plates in 20% fetal bovine serum (FBS) Dulbecco modified Eagle medium (DMEM) and then cultured for 24 hours in 0.5% FBS DMEM before exposure to either 1, 5, or 10 ng/mL of TGF-beta in serum-free DMEM for 12 or 24 hours or to 20, 80, or 300 ng/mL of PGE2 in serum-free DMEM for 12 or 24 hours. After incubation, cells were removed from plates by scraping and assayed for connexin-43 protein, first by Western blot to confirm the specificity of the anti-connexin-43 antibody and then by slot blot analysis for quantitative comparison of connexin-43 expression. Our studies showed no significant changes in connexin-43 expression in response to either factor. These studies suggest that exogenous TGF-beta and PGE2 do not alter connexin-43 expression.

Alveolar Process↗
PubMed2001

Relation between 9-aminocamptothecin systemic exposure and tumor response in human solid tumor xenografts.

9-Aminocamptothecin (9-AC) is a topoisomerase I inhibitor with activity against xenografts from childhood solid tumors; however, clinical trials with this compound have been disappointing, resulting in discontinuation of further development. The objectives of this study were to evaluate the antitumor activity of 9-AC in a panel of pediatric solid tumor xenografts and to relate the 9-AC lactone systemic exposure, defined as area under the concentration time curve (AUC), to the antitumor dose associated with tumor regression in the xenograft model. We evaluated protracted administration of i.v. and oral therapies (daily times 5) for 1, 2, or 3 weeks and for 1 or 3 cycles. The minimum effective dose of 9-AC causing objective regression of advanced tumors was determined for each schedule. 9-AC lactone plasma concentration-time profiles associated with the lowest dose achieving complete and partial responses for each xenograft were then determined for each regimen. Tumors were highly sensitive to 9-AC therapy, but the systemic exposure required for antitumor effect is in excess of that achievable in patients.

Animals↗
PubMed2000

Neuropsychological deficits in pediatric patients with childhood-onset schizophrenia and psychotic disorder not otherwise specified.

OBJECTIVE: Children with transient psychotic symptoms and serious emotional disturbances who do not meet current criteria for schizophrenia or other presently recognized diagnostic categories commonly present diagnostic and treatment problems. Clarifying the connections between children with narrowly defined schizophrenia and children with a more broadly defined phenotype (i.e., Psychotic Disorder Not Otherwise Specified, PD-NOS) has implications for understanding the pathophysiology of schizophrenia. In this study, the neuropsychological test performance of a subgroup of children with atypical psychosis was compared with that of patients with childhood-onset schizophrenia (COS). METHOD: Cognitive function was assessed with neuropsychological test battery regimens in 51 neuroleptic-nonresponsive patients within the first 270 at NIMH testing (24 PD-NOS, 27 COS) were included in this analysis. Seventeen (39%) of 44 COS subjects were unavailable for this study as their IQ tested <70. The PD-NOS patients were younger than the COS patients at the time of testing (12.0+/-2.8 vs 14.4+/-1.8years, respectively, p<0.004). The test levels of these groups were compared with each other. RESULTS: The neuropsychological test results for the PD-NOS and COS patients were 1-2standard deviations below normative data across a broad array of cognitive functions. There were no overall differences in the test levels for the six summary scales (F=2.82, df=1, 36, p=0.10) or in the profile shape (F=1.70, df=5, 180, p=0.14) between the PD-NOS and COS groups. For the COS patients, there was a significant difference between their mean full-scale WISC IQ (84.7+/-16.2) and their average standard scores for both the spelling (97.7+/-16.1, n=23, t=4.0, p=0.001) and reading decoding subtests (97.7+/-13.7, n=23, t=3.7, p=0.001) of the Kaufman Test of Educational Achievement. CONCLUSIONS: Treatment-refractory PD-NOS and COS patients share a similar pattern of generalized cognitive deficits, including deficits in attention, learning and abstraction which are commonly observed in adult patients with schizophrenia. These data support a hypothesis that at least some of the PD-NOS cases belong within the schizophrenic spectrum, which is of importance for future genetic studies planned for this cohort.

Adolescent↗
PubMed2000

Prompt and durable engraftment in two older adult patients with high risk chronic myelogenous leukemia (CML) using ex vivo expanded and unmanipulated unrelated umbilical cord blood.

Delayed engraftment, graft failure, and adverse transplant-related events have been observed in unrelated umbilical cord blood (UCB) recipients, particularly in those receiving a low leukocyte cell dose and in CML patients. We report the outcomes of two older adult patients with high risk CML who received a low leukocyte cell dose of unmanipulated UCB cells supplemented with ex vivo expanded (AastromReplicell System) UCB cells. Each engrafted promptly and neither patient experienced GVHD or life-threatening infection. Both remain engrafted with cells exclusively of donor origin and are in cytogenetic remission at 19 and 8 months follow-up. Ex vivo expanded UCB cells appear to facilitate hematopoietic recovery and therefore may increase the number of CML patients eligible for unrelated UCB transplant.

Antigens, CD↗
PubMed2000

Pharmacokinetics of irinotecan and its metabolites SN-38 and APC in children with recurrent solid tumors after protracted low-dose irinotecan.

Irinotecan (IRN), a topoisomerase I interactive agent, has significant antitumor activity in early Phase I studies in children with recurrent solid tumors. However, the disposition of IRN and its metabolites, SN-38 and APC, in children has not been reported. Children with solid tumors refractory to conventional therapy received IRN by a 1-h i.v. infusion at either 20, 24, or 29 mg/m2 daily for 5 consecutive days for 2 weeks. Serial blood samples were collected after doses 1 and 10 of the first course. IRN, SN-38, and APC lactone concentrations were determined by an isocratic high-performance liquid chromatography assay. A linear four-compartment model was fit simultaneously to the IRN, SN-38, and APC plasma concentration versus time data. Systemic clearance rate for IRN was 58.7 +/- 18.8 liters/h/m2 (mean +/- SD). The mean +/- SD ng/ml x h single-day lactone SN-38 area under the concentration-time curve (AUC(0-->6) was 90.9 +/- 96.4, 103.7 +/- 62.4, and 95.3 +/- 63.9 at IRN doses of 20, 24, and 29 mg/m2, respectively. The relative extent of IRN conversion to SN-38 and metabolism to APC measured after dose 1 were 0.49 +/- 0.33 and 0.29 +/- 0.17 (mean +/- SD). No statistically significant intrapatient difference was noted for SN-38 area under the concentration-time curve. Large interpatient variability in IRN and metabolite disposition was observed. The relative extent of conversion and the SN-38 systemic exposure achieved with this protracted schedule of administration were much greater than reported in adults or children receiving larger intermittent doses.

Adolescent↗
PubMed1999

Effects of sewage effluent discharge on the abundance, condition and mortality of hulafish, Trachinops taeniatus (Plesiopidae).

The effects of sewage effluent on the abundance, condition (length and reproduction) and mortality of hulafish Trachinops taeniatus were investigated at multiple outfall and control locations on the central coast of New South Wales, Australia. Underwater visual surveys found consistently fewer T. taeniatus at locations where sewage was discharged compared to control locations. The condition of T. taeniatus was investigated by comparing mean length and reproductive indices of fish from two outfall and two control locations. Fish from the largest outfall location were significantly smaller in comparison with fish from control locations. Gravid female fish from outfall locations had similar gonadosomatic indices but a significantly greater number of eggs and smaller size of eggs in comparison with fish from control locations. Mortality of T. taeniatus was investigated during 2-week, in situ, caging experiments at multiple locations and times and 80% of fish survived, although mortalities of up to 73% per cage were recorded at one sewage outfall. T. taeniatus may be a suitable environmental indicator species of sewage pollution.

Journal Article↗