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Biomedical subjects

K T Smith

Publications and source records attributed to K T Smith.

At least 19 recordsLinked to original sources
PubMed1996

Gene delivery systems for use in gene therapy: an overview of quality assurance and safety issues.

The development of safe and effective agents for gene therapy is founded on three main principles; careful choice and design of vectors, assessment of vector safety under GLP and production of the vector stocks under GMP. The first ensures the safe and appropriate contained delivery and expression of the required gene to the recipient of the therapy. GLP provides fully documented studies of potency, efficacy and safety of the product while the production of clinical grade agents under GMP is essential.

Genetic Therapy↗
PubMed1995

Plaque-associated expression of human herpesvirus 6 in multiple sclerosis.

Representational difference analysis was used to search for pathogens in multiple sclerosis brains. We detected a 341-nucleotide fragment that was 99.4% identical to the major DNA binding protein gene of human herpesvirus 6 (HHV-6). Examination of 86 brain specimens by PCR demonstrated that HHV-6 was present in > 70% of MS cases and controls and is thus a commensal virus of the human brain. By DNA sequencing, 36/37 viruses from MS cases and controls were typed as HHV-6 variant B group 2. Other herpesviruses, retroviruses, and measles virus were detected infrequently or not at all. HHV-6 expression was examined by immunocytochemistry with monoclonal antibodies against HHV-6 virion protein 101K and DNA binding protein p41. Nuclear staining of oligodendrocytes was observed in MS cases but not in controls, and in MS cases it was observed around plaques more frequently than in uninvolved white matter. MS cases showed prominent cytoplasmic staining of neurons in gray matter adjacent to plaques, although neurons expressing HHV-6 were also found in certain controls. Since destruction of oligodendrocytes is a hallmark of MS, these studies suggest an association of HHV-6 with the etiology or pathogenesis of MS.

Adult↗
PubMed1994

Detection, cloning and characterisation of papillomaviral DNA present in sarcoid tumours of Equus asinus.

Molecular investigations on 18 naturally occurring sarcoid tumors removed from donkeys identified papillomaviral DNA homologous to bovine papillomavirus (BPV)-2 DNA under stringent conditions, in all the samples. Restriction endonuclease analysis of 15 of the tumours demonstrated papillomaviral DNA similar to BPV-1 and BPV-2. The type of DNA was not specific to either the site or the type of lesion. The analysis of the nucleotide base sequence of a cloned papillomaviral element from a sarcoid showed that the isolate was 96 and 98 per cent homologous to BPV-1 in the L1 and E5 open reading frames, respectively. It was concluded that the disease in the donkey is similar to that in the horse and that the E5 open reading frame may be involved in oncogenesis in the sarcoid.

Animals↗
PubMed1994

Spinal bone loss in postmenopausal women supplemented with calcium and trace minerals.

The effects of calcium supplementation (as calcium citrate malate, 1000 mg elemental Ca/d) with and without the addition of zinc (15.0 mg/d), manganese (5.0 mg/d) and copper (2.5 mg/d) on spinal bone loss (L2-L4 vertebrae) was evaluated in healthy older postmenopausal women (n = 59, mean age 66 y) in a 2-y, double-blind, placebo-controlled trial. Changes (mean +/- SEM) in bone density were -3.53 +/- 1.24% (placebo), -1.89 +/- 1.40% (trace minerals only), -1.25 +/- 1.46% (calcium only) and 1.48 +/- 1.40% (calcium plus trace minerals). Bone loss relative to base-line value was significant (P = 0.0061) in the placebo group but not in the groups receiving trace minerals alone, calcium alone, or calcium plus trace minerals. The only significant group difference occurred between the placebo group and the group receiving calcium plus trace minerals (P = 0.0099). These data suggest that bone loss in calcium-supplemented, older postmenopausal women can be further arrested by concomitant increases in trace mineral intake.

Aged↗
PubMed1994

Evaluation of apheresis platelet concentrates collected with a reduced (30-ml) collection chamber with resuspension and storage in a synthetic medium.

Recently, the CS-3000 Plus Blood Cell Separator with the TNX-6 platelet separation chamber insert has been furnished with a small-volume (30-ml) collection chamber. In this study, a platelet synthetic medium containing glucose and bicarbonate (PSM) was used for resuspension and storage of this highly concentrated platelet product. Eighteen donors participated in a paired study design where each participant donated platelets on two occasions, once following collection in a standard chamber with resuspension and storage in plasma and once following collection in the new chamber with resuspension and storage in PSM. Substantially higher total platelet counts were obtained using platelets collected in the small chamber and stored in PSM as compared to control (4.4 +/- 0.9 x 10(11) vs. 3.5 +/- 0.9 x 10(11) platelets, p < 0.01 by paired t test). After 5 days of storage, PSM-stored platelets demonstrated higher ATP levels, less lactate dehydrogenase in the supernatant and increased lactate production with resulting lower pH at day 5 of storage (6.94 +/- 0.15 vs. 7.08 +/- 0.09, p < 0.05). There were no statistically significant differences of the survival by multiple-hit estimation of PSM-stored as compared to plasma-stored platelets as determined by 111In labeling and infusion. A slight decrease in the initial percent recovery with the additive-suspended as compared to suspended plasma cells was noted: 50 +/- 8 versus 54 +/- 9%, respectively (p < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1994

Timing of peak bone mass in Caucasian females and its implication for the prevention of osteoporosis. Inference from a cross-sectional model.

To determine the timing of peak bone mass and density, we conducted a cross-sectional study of bone mass measurements in 265 premenopausal Caucasian females, aged 8-50 yr. Bone mass and bone mineral density were measured using dual X-ray absorptiometry and single-photon absorptiometry at the spine (anteroposterior, lateral), proximal femur, radius shaft, distal forearm, and the whole body. Bone mass parameters were analyzed using a quadratic regression model and segmented regression models with quadratic-quadratic or quadratic-linear form. The results show that most of the bone mass at multiple skeletal locations will be accumulated by late adolescence. This is particularly notable for bone mineral density of the proximal femur and the vertebral body. Bone mass of the other regions of interest is either no different in women between the age of 18 yr and the menopause or it is maximal in 50-yr-old women, indicating slow but permanent bone accumulation continuing at some sites up to the time of menopause. This gain in bone mass in premenopausal adult women is probably the result of continuous periosteal expansion with age. Since rapid skeletal mineral acquisition at all sites occurs relatively early in life, the exogenous factors which might optimize peak bone mass need to be more precisely identified and characterized.

Adolescent↗
PubMed1993

Intraduodenal delivery of intrinsically and extrinsically labelled CaCO3 in the rat: effect of solubilization on calcium bioavailability.

The dissolution of CaCO3 before intraduodenal administration was found to be an important factor determining calcium (Ca) bioavailability. Extrinsically and intrinsically labelled 47CaCO3 preparations were sequentially dissolved by serial additions of HCl. Aliquots of these preparations were collected before (no HCl added) and during the solubilization process and administered intraduodenally to rats. Whole body 47Ca retention 72 h post-dose was used as a measure of Ca bioavailability. Although dissolution of CaCO3 significantly increased Ca bioavailability (P < 0.001), Ca from both intrinsically and extrinsically labelled CaCO3 was absorbed and retained to some extent without prior acid dissolution. Due to a disproportionately high concentration of 47Ca on the particle surface, extrinsically labelled 47CaCO3 overestimated bioavailability when unsolubilized or partially solubilized CaCO3 preparations were used (P < 0.05). These data indicate that dissolution is a determining factor for Ca bioavailability from CaCO3. Incomplete dissolution will significantly limit but not completely prevent Ca bioavailability. The disintegration and dissolution characteristics of commercial CaCO3 preparations, which vary widely, may produce important differences in Ca absorption.

Animals↗
PubMed1993

Effect of age, calcium source, and radiolabeling method on whole body 47Ca retention in the rat.

In a longitudinal study we determined the effect of animal age as well as Ca source and radiolabeling method on Ca bioavailability by measuring whole body 47Ca retention (WBR) in male Sprague-Dawley rats. The WBR assay was performed without surgery or anesthesia, and the same groups of animals were studied at 8, 16, 20, and 32 wk of age. Rats were administered a 6-mg radiolabeled oral dose of Ca as Ca citrate malate (CCM) or intrinsically or extrinsically labeled CaCO3 or hydroxyapatite (HAP). Fractional Ca retention was measured from the 72-h postdose WBR divided by WBR at time 0. WBR was significantly affected by Ca source with CCM > CaCO3 > HAP at all ages (P < 0.001). The rank order and relative bioavailabilities of these Ca salts in the rat model agreed well with literature values for Ca absorption in adult humans. Although percent WBR decreased significantly with advancing age (P < 0.001), the mean rate of decline (-3.4%/wk) was not affected by Ca source. Extrinsic radiolabeling overestimated (approximately 20%) Ca bioavailability when the rats were young. However, the magnitude of this effect diminished with advancing animal age and was not significant across all ages (repeated measures analysis of variance P = 0.10).

Administration, Oral↗
PubMed1993

BPV-4 induces amplification and activation of 'silent' BPV-1 in a sub-line of C127 cells.

BPV-4-induced malignant transformation of C127 mouse fibroblasts in vitro is the result of a 'hit and run' mechanism. Viral DNA is lost on continued subculture of transformed cell lines without loss of any malignant characteristics. The DNA from these cells harbours specific amplified sequences. Two such amplified fragments of approximately 10 kb and 12 kb were molecularly cloned and designated HL-10 and HL-12 respectively. HL-10 transformed C127 cells efficiently and therefore encodes a transforming function, whereas the 12 kb clone did not. Heteroduplexes showed that HL-12 was homologous to HL-10 except for two additional tandem copies of an approximately 1.7 kb sequence. Sequence analysis of HL-10 revealed that the clone contained a 5.2 kb region from BPV-1 including the transforming ORFs. Transformation studies have shown differences between HL-10 and BPV-1, indicating that the host flanking sequences may contribute to the transforming potential of the BPV-1 ORFs. The BPV-1 DNA was associated with sequences homologous to murine autonomously replicating sequences (ARS) implicated in the establishment of multiple tandem DNA repeats. As the parental cells contain the set of sequences amplified in transformed cells in single copy and show none of the characteristics of transformed cells, we conclude that BPV-4 has activated these sequences by amplification and rearrangement. These phenomena may be mediated through an interaction between BPV-4 proteins and the BPV-1 origin of DNA replication or via the ARS region.

Animals↗
PubMed1992

Association of bovine papillomavirus type 2 and bracken fern with bladder cancer in cattle.

The bladder cancer syndrome that often accompanies chronic enzootic hematuria in cattle grazing on pastures infested by bracken fern has been experimentally reproduced in animals fed a diet of bracken. The experimentally induced tumors were histologically and pathologically indistinguishable from the naturally occurring ones and comprised two main types: (a) carcinoma of the urothelium identical to that seen in humans; and (b) hemangioendotheliomas of the subjacent capillaries. Often the two types of tumor occurred together in the same bladder. In animals experimentally immunosuppressed with azathioprine "bracken type" hemangiomas developed in the bladder lining. DNA of bovine papillomavirus (BPV) type 2 was found in 46% (7 of 15) of the natural cancer cases and in 69% (9 of 13) of the experimentally induced lesions, independently of histological type and including the hemangiomas of the azathioprine-treated animals, suggesting a close association between BPV and bovine bladder neoplasia. Moreover, BPV-2 DNA was found in experimental animals that had not been inoculated with BPV at all or had been inoculated with a different BPV type and had been kept in isolation, suggesting that BPV can persist in a latent state and be activated when the animal is exposed to the bracken cocarcinogens and to immunosuppressants.

Animals↗
PubMed1991

The B subgroup bovine papillomaviruses lack an identifiable E6 open reading frame.

Analysis of the corrected DNA sequence for the bovine papillomavirus type 4 (BPV4) genome revealed that there is no open reading frame (ORF) that might encode an E6 protein. The other two B subgroup bovine papillomaviruses, BPV3 and BPV6, were found to have the same arrangement of ORFs in this region as BPV4. Thus, we conclude that E6 functions are either not required by these viruses or are performed by another viral (or host) protein. Furthermore, the position that might be expected to be occupied by E6, between the long control region and the E7 ORF, contains the E8 ORF, which has the potential to encode a 42-residue polypeptide with considerable similarity to the E5 transforming protein of BPV1. Therefore, it appears that during the evolution of the B subgroup of BPVs, genomic rearrangements may have occurred resulting in the present layout of the early ORFs.

Amino Acid Sequence↗
PubMed1991

Studies on vaccination against papillomaviruses: prophylactic and therapeutic vaccination with recombinant structural proteins.

The L1 and L2 proteins of BPV-2 have been produced in Escherichia coli as beta-galactosidase fusion proteins. The fusion proteins have been used to vaccinate calves both prophylactically and therapeutically. The L1 fusion protein prevented tumor formation when administered before challenge with BPV-2, while the L2 fusion protein was very effective in promoting tumor rejection, independently from whether it was administered before or after challenge. Animals vaccinated with L1, but not with L2, responded rapidly with production of serum neutralizing antibodies, showing that this peptide contains B-cell-specific epitopes. The massive infiltration of lymphocytes in the tumors of L2-vaccinated animals suggests that the peptide contains epitopes specific for T-cells. The two structural proteins of BPV-2 therefore interact with both efferent arms of the immune system, and this observation allows the choice between two different types of antiviral vaccination.

Animals↗
PubMed1991

Spinal bone density and calcium intake in healthy postmenopausal women.

Dietary calcium intake and bone mineral density (BMD) of the lumbar spine (L2-L4) were determined in 131 healthy free-living postmenopausal women (aged 64.7 +/- 7.6 y, means +/- SD). The calcium consumption for the total population was 606 +/- 302 mg/d. Subjects consuming less than the population mean of dietary calcium had significantly lower BMDs than did subjects with intakes above the mean (P less than 0.009); these two groups did not differ in basic demographic characteristics. Additional analyses using a stepwise univariate regression model demonstrated that BMD was significantly associated with body weight (P less than 0.001) and dietary calcium intake (P less than 0.02). These data support the hypothesis that dietary calcium intake is a determinant of skeletal health in postmenopausal women.

Aged↗
PubMed1991

Malignant transformation of a papilloma induced by bovine papillomavirus type 4 in the nude mouse renal capsule.

A papillomatous cyst was induced by implanting bovine foetal palate epithelium, infected in vitro with bovine papillomavirus type 4 (BPV-4), beneath the renal capsule of a nude mouse. The benign tumour underwent malignant progression, developing into a squamous cell carcinoma with metastatic deposits in the spleen. The bovine origin of both the renal and splenic cancers was confirmed by the presence of bovine major histocompatibility complex class I antigens in the cancer cells and by sequencing the Harvey-ras 1 gene, which was shown to be of bovine origin. BPV-4 DNA was present in the residual papillomatous fronds of the renal cancer, but was absent from the carcinoma proper and for the splenic metastasis. These results confirm that BPV-4 is a carcinogenic agent and that its genetic information is not necessary for the maintenance of the malignant phenotype. Moreover the system provides the opportunity to investigate the role of viral and chemical carcinogens in an experimental system.

Animals↗
PubMed1990

Studies on vaccination against papillomaviruses: the immunity after infection and vaccination with bovine papillomaviruses of different types.

Calves, free of antibodies to bovine papillomaviruses (BPV), were reared in isolation. One was infected with BPV-2, developed tumours and was resistant to homologous reinfection. Groups of calves were infected with BPV-2, BPV-5 or BPV-6; they all developed and subsequently rejected type-specific tumours. They were then infected with BPV-4; they were not immune and oral papillomas were induced. Groups of animals were vaccinated by intramuscular preparations of purified BPV-4 and BPV-6 and were challenged with homologous virus; all were immune to reinfection. An earlier experiment had shown this to be true for BPV-2. Two calves, immune to BPV-6, were not immune to BPV-1. These experiments, although they do not cover all the possibilities of reciprocal immunisation and challenge, indicate that prophylactic immunity to a range of papillomaviruses is type-specific. This is the first clear demonstration of this phenomenon in the papillomavirus group.

Animals↗
PubMed1990

Studies on vaccination against papillomaviruses: a comparison of purified virus, tumour extract and transformed cells in prophylactic vaccination.

Calves were vaccinated with two preparations made from one cutaneous fibropapilloma induced by bovine papillomavirus type 2 (BPV-2). One vaccine consisted of homogenised tumour; the other contained purified virus only. Both produced resistance to a heavy challenge infection of BPV-2. One calf in the vaccinated group developed a small tumour and rejected it earlier than the control calves. It would appear likely that the prophylactic immune response was induced by viral structural proteins only and that tumour-specific antigens are unnecessary. Bovine fibroblasts were transformed in vitro by BPV-2 and administered as a vaccine; immunity was not induced.

Animals↗