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Biomedical subjects

R A Roberts

Publications and source records attributed to R A Roberts.

At least 91 records · Page 5Linked to original sources
PubMed1989

Activation of the cellular Harvey ras gene in mouse skin tumors initiated with urethane.

Mouse skin tumors, benign papillomas, and squamous cell carcinomas (SCCs) were initiated by a single topical application of urethane followed by repeated promotion with 12-O-tetradecanoylphorbol-13-acetate (TPA). Using the NIH 3T3 focus forming assay, dominant transforming activity was detected in DNA isolated from SCC samples. Rearranged and amplified copies of the c-Ha-ras gene were detected in NIH 3T3 transformant cell lines, indicating that an activated Ha-ras gene had been transferred to the NIH 3T3 recipient cells. Analysis of p21ras from the transformant cell lines suggested that the activating ras mutation was present in codon 61. Ultimately, the Ha-ras gene was shown to be activated by a specific A----T transversion at the second position of codon 61. This mutation was detected in both benign papillomas and SCCs, suggesting the activation occurred early in tumor development. The results demonstrate a highly consistent activation of the Ha-ras oncogene by a specific point mutation, suggesting a functional role for an activated ras gene in the initiation of mouse skin tumors by urethane.

Animals↗
PubMed1989

Bradykinin receptors: characterization, distribution and mechanisms of signal transduction.

Bradykinin is a peptide consisting of nine amino acids. It is a member of the kinin family, a class of molecules sometimes considered to be locally acting hormones. Bradykinin acts through cell surface receptors to elicit a series of biological responses, many of which have been well characterized at the whole organ or body level. However, little is known about the bradykinin receptor itself or its mechanisms of signal transduction, its function and its tissue distribution. Increasing evidence suggests that bradykinin is a member of a group of locally produced peptides which may act in a paracrine fashion as microenvironmental modulators of cell proliferation. Evidence for this derives from studies of the interaction between bradykinin and its receptor, receptor-effector coupling systems and in vitro studies of the biological effects of bradykinin. These areas, together with questions concerning the nature and number of different types of bradykinin receptors, form the main bulk of current interest in bradykinin research and are the subject of this review. The ability of bradykinin to synergize with other growth regulating ligands will also be considered.

Amino Acid Sequence↗
PubMed1989

Bradykinin receptor number and sensitivity to ligand stimulation of mitogenesis is increased by expression of a mutant ras oncogene.

Bradykinin is a nine amino acid peptide of the kinin family believed to play a role in pain mediation and in the regulation of blood pressure, fluid balance and smooth muscle contraction. Here we demonstrate that bradykinin is also a potent mitogen for a mutant Ha-ras-transfected cell line, Rat 13. The Rat 13 cells display two binding sites for bradykinin: a moderate number (52,000) of high affinity sites (Kd = 4.9 nM) co-exist with a much smaller number (1100) of very high-affinity sites (Kd = 2.7 pM). Ligand binding stimulates mitogenesis through the lower affinity receptors, which are classified as B2. These receptors are down-regulated in response to ligand. In contrast, Rat 1 cells (the cell line from which Rat 13 was derived) have only 4000 receptors per cell in total and respond weakly to bradykinin as a mitogen. Thus, expression of a mutant ras protein in Rat 13 cells increases their expression of the bradykinin receptor and their sensitivity to ligand stimulation of mitogenesis. Additional binding studies demonstrate that human A431 epithelial cells and Swiss mouse 3T3 fibroblasts also possess high-affinity sites for bradykinin.

Animals↗
PubMed1988

Protease inhibitors decrease rabbit cartilage degradation after meniscectomy.

In vitro proteoglycan (PG) synthesis and release were measured on cartilage removed from rabbit knees within 1 week of meniscectomy. Three days following partial lateral meniscectomy, 72% of the femurs and 82% of the tibias had visible ulcers. Cartilage from the weight-bearing areas incorporated 2.0-2.9 times more 35S-sulfate in vitro than cartilage from the opposite, unoperated knees. 3H-thymidine incorporation was 2.5-3.4 times higher for surgical than control groups. 35S-sulfate incorporation by the surgical group was inhibited by 22% in the presence of 10(-4) M U24522, an inhibitor of rabbit chondrocyte metalloprotease (CMP). 3H-thymidine incorporation by the surgical group was inhibited by 28% by 10(-4) M U24522. In vitro PG release from cartilage removed 2 days after surgery was 1.6-3.7 times higher for the surgical than the control group. PG release by the surgical group after 22 h of incubation was reduced to the control level by three CMP inhibitors, U24278, U24279, and U24522. PG release by cartilage from the nonsurgical group was also reduced by these compounds at 22 h. These results suggest that both the anabolic and catabolic processes that are stimulated by surgery can be isolated in vitro and that CMP may be involved in the catabolic process.

Animals↗
PubMed1987

Proteoglycan degradation by a chondrocyte metalloprotease. Effects of synthetic protease inhibitors.

Synthetic inhibitors of a chondrocyte metalloprotease (CMP) were assessed for potency. Proteoglycan core protein was used as substrate. The IC50 values were between 2 X 10(-6) and 7 X 10(-6) M for two types of inhibitors, thiol tripeptides and N-carboxyalkyl peptides. Hydroxamic acid peptides were more potent, with IC50 values of 3.2 X 10(-8) to 6.0 X 10(-8) M. These results confirm inhibitory concentrations reported using a proteoglycan-polyacrylamide bead assay. The slopes of the dose-response curves for the thiol compounds were steeper than the slopes for the other two types of compounds. All of the culture media tested inhibited CMP to some extent. Some media also interfered with inhibitor activity. In Ham's F10 nutrient medium, minimum CMP inhibition occurred, and all four hydroxamic acid peptides retained their activity for 1-2 days at 37 degrees. One thiol peptide compound assayed lost activity in 1 hr in thiocyanate-treated serum. All four hydroxamic acid peptides assayed retained activity in thiocyanate-treated serum after 3 days at 37 degrees. The hydroxamic acid peptides may provide a way to block endogenous CMP activity in vivo and to assess the role of CMP in normal and experimentally altered cartilage. They are more potent than other known CMP inhibitors. They retain activity in culture media and serum conditions used for in vivo and in vitro tests of CMP activity and toxicity.

Animals↗
PubMed1987

Metabolically inactive 3T3 cells can substitute for marrow stromal cells to promote the proliferation and development of multipotent haemopoietic stem cells.

When highly enriched multipotential spleen colony forming cells (CFU-S) obtained following fluorescence activated cell sorting (FACS-CFU-S) are cultured on marrow stromal cells, they undergo proliferation and development to produce mature haemopoietic cells (Spooncer et al., Nature, 316:62-64, 1985). We now show that FACS-CFU-S behave in a similar way when cultured on monolayers of 3T3 cells, indicating that the 3T3 cells can supply at least part of the environment which is representative of marrow stromal cells and provide, therefore, a system for studying stromal cell: haemopoietic cell interactions. We also demonstrate that IL-3-dependent multipotential stem cell lines (FDCP-Mix), but not a variety of other "committed" IL-3-dependent cell lines, resemble FACS-CFU-S in terms of their ability to proliferate and differentiate when cultured on 3T3 cells in the absence of IL-3. In this system, attachment of the FDCP-Mix to the 3T3 cells is critical for the subsequent maintenance of viability and stimulation of development of the cells. When the FDCP-Mix cells are physically separated from the 3T3 cells, they die and their death cannot be prevented by using 3T3-cell-conditioned medium. The extracellular matrix generated by 3T3 cells is not sufficient for promoting attachment or viability of the FDCP-Mix cells, indicating the importance of integral membrane components. However, attachment and development of FDCP-Mix cells occurs on 3T3 cells that have been lightly fixed with glutaraldehyde indicating that active metabolism is not essential for the effects promoted by the 3T3 cells. We suggest that the ability of FACS-CFU-S and FDCP-Mix cells to respond to 3T3 cells involves specific ligand/receptor interactions.

Animals↗
PubMed1987

Effect of synthetic metalloprotease inhibitors on cartilage autolysis in vitro.

The role of chondrocyte metalloprotease (CMP) in mediating cartilage autolysis was studied. Proteoglycan (PG) release and synthesis by rabbit articular cartilage explants were measured. After a 1-day preculture in control medium, 3.3 X 10(-6) M retinoic acid (RET) treatment for 1 day stimulated PG release several fold. RET also caused a large decrease in PG synthesis that returned to the control level after a 3-day recovery period. The effect on PG synthesis was observed at serum levels of 5 and 0.05%. The effect of RET on PG release required protein synthesis, inasmuch as it was lost in cultures maintained in media without amino acids or in a low volume of media. Interleukin-1 (IL-1) and lipopolysaccharide (LPS) treatment for 2 days also stimulated PG release. More PG was released after RET than after IL-1 or LPS, and only RET produced an effect that was evident by day 1. The amount of CMP that produced the same size effect on PG release as these stimulators was below the detection level of PG protease assays. Three potent CMP inhibitors reduced RET-, IL-1- and LPS-stimulated PG release to control levels. These inhibitors did not block another action of RET on chondrocytes, namely the inhibition of PG synthesis by RET immediately after treatment. The inhibitors did not act by reducing cell viability, because recovery of the rate of PG synthesis 3 days post-treatment occurred in inhibitor-treated cultures. These studies suggest that CMP is involved in cartilage autolysis that is stimulated by RET and IL-1.

Animals↗
PubMed1987

Conservative management of renal lacerations in blunt trauma.

Controversy in the treatment of blunt renal trauma is largely focused on immediate surgery versus conservative management for parenchymal lacerations. A retrospective analysis of 133 cases of blunt renal trauma at the Victoria General Hospital in Halifax over a 10-year period revealed 26 cases of renal laceration. The conservative approach to radiologic diagnosis and treatment options is discussed. Our experience confirms a low rate of both nephrectomy and secondary complications using conservative management.

Accidents, Traffic↗
PubMed1986

Suction socket suspension for below-knee amputees.

In this study the current use of suction suspension for below-knee prostheses is examined by means of two questionnaire surveys. The experience of 56 below-knee (B-K) amputees wearing suction socket prostheses is evaluated comparing suction prostheses with previously worn limbs. A high degree of satisfaction was found, with amputees on the whole reporting improved skin condition, diminished pain, and increased activity levels compared to previous prosthetic history. The experience and opinions of 466 certified prosthetist members of the American Orthotist Prosthetist Association are examined in the second survey, including degree of contact, success, and evaluation of problems in using suction suspension for the B-K amputee. This survey indicated limited contact and familiarity with B-K suction suspension, with only 22% stating they had made this type of prosthesis. Prosthetists cited characteristics of the B-K residual limb as the chief deterrent to a successful fitting.

Adult↗
PubMed1986

The University of California Biomechanics Laboratory four-bar polycentric knee linkage. A clinical trial in 20 active above-knee amputees.

A four-bar polycentric prosthetic knee linkage developed by the University of California Biomechanics Laboratory (UCBL) was tested clinically in 20 active veteran above-knee amputees. The mean follow-up period was 12.3 months. At final follow-up evaluation, positive subjective responses were reported by 80% of the amputees. More than one-third reported that the device was superior to any that they had used previously. Almost all subjects found that prior levels of daily activity and the ability to perform specific activities were maintained or enhanced. The increase in knee flexion afforded by the mechanism was found to be an advantage. There was no significant change in various fundamental gait parameters. Problems caused by excessive wear of anodized aluminum linkage pins and by air leaks in the pneumatic swing-phase control unit were corrected easily. These preliminary results encourage further clinical application and study of the UCBL four-bar polycentric knee.

Adult↗
PubMed1985

Subjective evaluation of the UCBL polycentric knee linkage by active above knee amputees. A prospective trial.

In this study 20 active male above knee amputees were fitted with prostheses incorporating prototype UCBL four-bar polycentric knee units. The subjective responses of the amputees were evaluated in a prospective fashion. Additional information concerning the short-term durability and mechanical behavior of the device is presented. The average followup was 12.3 months. Sixteen of the 20 subjects felt that they benefited from the device, and seven of these stated that the research prosthesis was superior to all previous devices that they had used. Four subjects stated that they had received little or no benefit from the research limb. Previous levels of function and the ease of performance of specific activities were maintained or enhanced in almost all subjects. Most subjects felt that the ease of walking on inclines was improved, and found the increase in knee flexion afforded by the mechanism to be an advantage. Early in the study, problems were encountered due to air leaks in the pneumatic swing-phase control units and to excessive wear of the aluminum linkage pins. These problems were corrected easily, and the devices were found to function well during the remainder of the study. Given the encouraging results from this preliminary clinical trial, we conclude that further study and clinical application of the UCBL four-bar polycentric knee are indicated.

Adult↗
PubMed1984

Reduced bisantrene-induced cytotoxicity and protein-associated DNA strand breaks under hypoxic condition.

The effect of hypoxia on bisantrene-induced cytotoxicity and protein-associated DNA strand breaks was studied in mouse L1210 leukemia cells, human T47D breast cancer cell line, and fresh samples of human tumor cells. For each cell type, there was enhanced cell killing under oxic versus hypoxic conditions (1-hour drug treatment at 37 degrees C). Likewise, in L1210 cells, the frequency of bisantrene (10 micrograms/ml for 1 hour)-induced protein-associated DNA strand breaks was reduced by 70% under hypoxic versus oxic conditions. The reason for enhanced cytotoxicity and DNA strand breakage in L1210 cells under oxic conditions could not be attributed to oxygen-enhanced cellular uptake of the drug. The clinical implication of these data is that bisantrene may be relatively ineffective against large tumors containing hypoxic cell populations.

Animals↗