Purification of cartilage-derived growth factor by heparin affinity chromatography

Cartilage-derived growth factor (CDGF) was found to bind tightly to columns of immobilized heparin and could be eluted with concentrations of salt in the order of 1.6-1.8 M NaCl. The molecular weight of CDGF was estimated to be 18,000-20,000 by high performance liquid-size exclusion chromatography. The affinity of CDGF for heparin greatly facilitated its purification. Highly purified CDGF active at about 1-2 ng/ml was obtained when crude cartilage extract was applied to heparin-Sepharose and the growth factor activity was recycled over heparin-Sepharose two more times. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver stain visualization of highly purified CDGF showed one major polypeptide band with a molecular weight of about 19,000 containing over 95% of the protein and one minor polypeptide band containing the rest of the protein. Only the Mr 19,000 polypeptide was active after elution from the polyacrylamide gel. Although CDGF bound tightly to immobilized heparin, it did not bind to immobilized chondroitin sulfate or hyaluronic acid. In addition, CDGF bound to heparin much more tightly than did platelet-derived growth factor even though these two growth factors had similar isoelectric points of about 10. These results suggest that the binding of CDGF to heparin was due to a specific affinity of the 2 molecules for each other.     

Herpesvirus sylvilagus infects both B and T lymphocytes in vivo.

Herpesvirus sylvilagus infection of cottontail rabbits (Sylvilagus floridanus) was studied as a model of herpesvirus-induced lymphoproliferative disorders. Leukocytosis, splenomegaly, proliferation of T cells and virus production by lymphocytes characterized this infectious mononucleosis-like disease. Approximately two copies of circular herpesvirus sylvilagus genomes per cell were detected in spleen cells at 2 weeks postinfection, and circular genomes could still be observed after 4 months. Circular viral genomes were found in both B and T lymphocytes. Small amounts of linear viral DNA (0.1 to 0.3 copies per cell) were also detected in both B and T cells. These results indicated that the virus did not replicate in the majority of lymphocytes in vivo. Herpesvirus sylvilagus infection in cottontail rabbits could be useful as a model for studying the complex virus-host relationships of lymphotropic herpesviruses and perhaps as an animal model for Epstein-Barr virus infection in humans.     

Effects of calmodulin and lipoxygenase inhibitors on LH (lutropin)- and LHRH (luliberin)-agonist-stimulated steroidogenesis in rat Leydig cells.

The results of this study, carried out with purified rat Leydig cells, indicate that there are no major differences in the stimulating effects of lutropin (LH) and luliberin (LHRH) agonists on steroidogenesis via mechanisms that are dependent on Ca2+. This was demonstrated by using inhibitors of calmodulin and the lipoxygenase pathways of arachidonic acid metabolism. All three calmodulin inhibitors used (calmidazolium, trifluoperazine and chlorpromazine) were shown to block LH- and LHRH-agonist-stimulated steroidogenesis. This probably occurred at the step of cholesterol transport to the mitochondria. Similarly, three lipoxygenase inhibitors (nordihydroguaiaretic acid, BW755c and benoxaprofen), inhibited both LH- and LHRH-agonist-stimulated steroidogenesis. The amounts of the inhibitors required were similar for LH- and LHRH-agonist-stimulated steroidogenesis. Steroidogenesis stimulated by the Ca2+ ionophore A23187 was also inhibited, but higher concentrations of the inhibitors were required. Indomethacin (a cyclo-oxygenase inhibitor) increased LHRH-agonist-stimulated steroidogenesis;this is consistent with the role of the products of arachidonic acid metabolism via the alternative, lipoxygenase, pathway. The potentiation of LH-stimulated testosterone production by LHRH agonist was unaffected by indomethacin or by lipoxygenase inhibitors at concentrations that inhibited LH-stimulated testosterone production by 75-100%. It was not possible to eliminate a role of calmodulin in modulating the potentiation, although higher concentrations of the inhibitors were generally required to negate the potentiation than to inhibit LH- or LHRH-agonist-stimulated testosterone production.     

Overlapping functional units in a cell division gene cluster in Escherichia coli

The ftsZ ( sulB ) coding sequence is preceded by two promoters, at least one of which lies within the coding sequence of the neighboring gene, ftsA . This region of the ftsA gene is required for full biological activity of ftsZ .     

In what sense is contemporary medicine dualistic?

Medicine's inability to humanely respond to the concerns of its patients has often been attributed to its Cartesian dualism of mind and body. More recently, this inability has been ascribed to medicine's penchant for isolating biological disease as its sole proper concern to the exclusion of experienced illness. This dualism of disease and illness is claimed to be an outgrowth of the Cartesian dualism but the differences and similiarities between these two forms of dualism is not clear. This paper seeks to clarify their relationship through an examination of their historical origins.Disease is currently identified and characterized by a process of clinico-pathological correlation. By this means clinical impressions are corrected in light of autopsy findings. Our current mode of clinico-pathological correlation originated in Paris in the early nineteenth century with the work of Xavier Bichat and others. The theory of knowledge implicit within this clinico-pathological approach to the body is described and compared to that of Descartes. Though medicine does pursue certain Cartesian goals for knowledge, such as knowledge of the patient that does not rely upon his candor, it ultimately espouses neither a Cartesian theory of knowledge nor a Cartesian theory of the body. With pathological inquiry modeled after the autopsy as the final word in the identification and characterization of disease, the patient's capacity for self-knowledge and interpretation — not an entity called mind — is isolated away from his body. This approach to the body means that objective evidence of disease is valued to the exclusion of subjective evidence.If the shortcomings of modern biomedicine are to be effectively addressed, not only must the interdependence of disease and illness be acknowledged. The very canons of medical evidence must be revised. Subjective evidence must be rehabilitated and rejuvenated with better methods of subjective clinical investigation. Host factors relevant to the healing and knowing of sickness must be elucidated so that medicine may rediscover the sentience of its patients. Department of Psychiatry and Behavioral Sciences University of Washington Seattle, WA 98195, U.S.A.     

The structure of cruciforms in supercoiled DNA: probing the single-stranded character of nucleotide bases with bisulphite.

The single-stranded character of cytosine bases in three cruciform structures has been assessed by an examination of reactivity towards sodium bisulphite. Unpaired cytosine residues undergo deamination at C4 to give deoxyuracil, and propagation in an ung Escherichia coli host results in C-G----T-A transition mutations, detectable by restriction cleavage or sequence analysis. Very high frequencies of such mutations have been found at cruciform loops, confirming their unpaired character, with almost zero background mutation frequencies elsewhere. A low level of modification was observed at the four-way junction of a cruciform. The results indicate that the optimal cruciform loop size is four bases, with loose 'breathing' at the first base pair at the top of the cruciform stem at 37 degrees C, and little or no opening of base pairs at the four-way junction.     

Stability of tissue culture medium pH as a function of autoclaving,time, and cultured plant material

Autoclaving is a standard procedure for sterilizing nutrient media for plant tissue cultures. Most tissue cultures are grown at pH 5.2 to 5.8 with pH adjustments being made prior to autoclaving. This paper reports that there are significant differences between initial pH levels and pH levels following autoclaving, particularly in the pH range of 5.7 to 8.5. This effect is noted with and without agar. In addition, we report that with time the pH of the medium drifts into the acid range. When Cucumis callus was added to the medium, the pH was changed significantly within 48 hours. The amount and direction (increase or decrease of pH) was significantly correlated with the original pH. This suggests that researchers should be wary of the true pH situation in their medium. In addition, in publications authors should specify whether their medium pH value was determined before or after autoclaving.     

Comparison of the Limulus amoebocyte lysate test with plate counts and chemical analyses for assessment of the quality of lean fish.

The quality of lean fish was assessed simply and rapidly with Limulus amoebocyte lysate. The endotoxin levels agreed with aerobic plate counts and chemical indices of spoilage. Correlation between level of endotoxin and level of total volatile bases was found to be highly significant (r = 0.8579; P less than 0.001).