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Biomedical subjects

G DiPasquale

Publications and source records attributed to G DiPasquale.

At least 19 recordsLinked to original sources
PubMed1991

Pharmacological modulation of rat monocytes: in vivo effects on Ia expression and interleukin-1 production.

We have shown that during the developing phase of adjuvant disease (AD) in rats the expression of MHC class II (Ia) antigens on blood monocytes (BM) was enhanced. The results of a study in established AD are reported now. Four agents were tested: indomethacin and diclofenac-sodium (1 mg/kg/day); levamisole and prinomide (10 mg/kg/day), administered orally from day 18-31 after induction of AD. We assessed the following BM parameters: Ia expression, interleukin-1 (sIL-1) production, and membrane bound IL-1 (mIL-1). In AD Ia expression was enhanced, no changes occurred in mIL-1 or sIL-1. Indomethacin treatment reduced sIL-1 production, levamisole Ia expression and mIL-1 activity, prinomide all three parameters measured and diclofenac, though clinically effective, none.

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

Interleukin-1 induces chondrocyte protease production: the development of collagenase inhibitors.

Supernatants from the P388D1 macrophage cell line as well as human interleukin-1 (IL-1) stimulated primary rabbit articular chondrocytes to produce collagen- (C-ase) and proteoglycan- (PG-ase) degrading proteases. The P388D1 derived factor had a molecular weight of 16,000-20,000 and a pI of 4.5-5.0. Both protease activities were metal dependent and inhibited by EDTA, phenanthroline, and alpha 2-macroglobulin but not by PMSF, TLCK, pepstatin, or alpha 1-antitrypsin. Size exclusion chromatography indicated the molecular weights for latent PG-ase and C-ase were 44,000-56,000 and 34,000-44,000, respectively. Chemical synthesis efforts produced two classes of C-ase inhibitors--thiols and hydroxamic acids. The former had IC50 values of 10(-5)-10(-6) M while the latter approached 10(-7) M.

Animals↗
PubMed1987

The potential use of implanted radiolabelled bovine nasal cartilage in dialysis tubing to evaluate agents affecting cartilage degradation in vivo.

Cartilage which undergoes extensive autolysis in vitro (spontaneous or stimulated) is characterized by proteoglycan loss. Experimental conditions and inhibitor profils studies suggest neutral metalloproteinases induce the autolysis. In these preliminary studies we compared the degradation of Na2 35SO4 labeled bovine nasal cartilage (BNC) plugs placed in dialysis tubing in vitro and in vivo. The dialysis tubing was used to exclude large molecules (molecular weights greater than 2000) like proteinases, and proteinase inhibitors (e.g. alpha 2-macroglobulin) but not potential test agents from the implanted cartilage. Cartilage autolysis occurred with live tissue but not with heat-killed tissue in both the in vitro and in vivo systems. In addition retinoic acid and phenanthroline were effective when placed inside or outside the dialysis tubing. A potentially useful procedure to evaluate agents which affect cartilage degradation is described.

Animals↗
PubMed1987

Progesterone receptor of adult rabbit lung.

Properties of progestin binding sites in adult male rabbit lung cytosol were analyzed using [3H] promegestone ([3H]R5020). At concentrations of 0.05-10 nM, [3H]R5020 bound to two saturable sites with differing affinities: a high affinity site (Kd = 0.34 nM, Bmax = 57 fmol/mg protein) and a moderate affinity site (Kd = 60 nM, Bmax = 540 fmol/mg protein). [3H]R5020 binding, under conditions selected to favor binding to the high affinity site, was reversible, protease-sensitive and inhibited in a concentration-dependent manner by steroids with the following order of potency: R5020 greater than norgestrel greater than norethindrone greater than progesterone greater than norethindrone acetate greater than norethynodrel greater than deoxycorticosterone greater than lynestrenol greater than quingestanol greater than testosterone greater than 17 beta-estradiol greater than cortisol. Upon sucrose density gradient ultracentrifugation analysis, a peak of [3H]R5020 binding activity was observed in the 6-7S region in the absence of KCl and in the 3-4S region in the presence of KCl. The progestin-binding activity of adult male rabbit lung cytosol thus possesses those characteristics conventionally accorded to a hormone receptor and suggests that lung tissue may be progesterone-responsive.

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↗
PubMed1986

Interleukin-1 stimulates the secretion of proteoglycan- and collagen-degrading proteases by rabbit articular chondrocytes.

Supernatants from the P388D1 murine macrophage cell line as well as commercially prepared human interleukin-1 (IL-1) stimulated primary rabbit articular chondrocytes to produce collagen- and proteoglycan-degrading proteases. The P388D1-derived factor had a molecular weight of 16,000-20,000 and a pI of 4.5-5.0, and was sensitive to phenylglyoxal treatment. Human IL-1 and the P388D1 supernatants enhanced glycosaminoglycan (GAG) release from bovine nasal cartilage explants. The proteoglycan- and collagen-degrading proteases required Ca2+ for activity. Latent proteoglycanase and collagenase had molecular weights of 44,000-56,500 and 34,000-44,000, respectively. The activated proteases had molecular weights of 30,000-40,000 and 22,000-36,000, respectively. Heparin-Sepharose affinity chromatography yielded two latent proteoglycanase-degrading protease activities and a collagen-degrading peak. The two proteoglycanase peaks also degraded fibronectin, laminin, gelatin, and azocoll but not type I collagen. The collagenase peak also degraded proteoglycan, gelatin, fibronectin, laminin, and azocoll. The activity of the proteoglycan- and collagen-degrading peaks was inhibited by phenanthroline and alpha 2-macroglobulin but not by phenylmethylsulfonylfluoride (PMSF), tosyllysylchloromethylketone (TLCK), pepstatin, or alpha 1-antitrypsin. The control of factors which augment protease production may offer a novel therapeutic approach to arthritis.

Animals↗
PubMed1986

Proteoglycan- and collagen-degrading enzymes from human interleukin 1-stimulated chondrocytes from several species: proteoglycanase and collagenase inhibitors as potentially new disease-modifying antiarthritic agents.

Human IL-1-stimulated chondrocytes derived from rabbit, bovine, and human articular cartilage produce proteoglycan- and collagen-degrading enzymes. These studies demonstrate that the biological activity of IL-1 is not species specific. Several thiol, carboxyalkyl, and hydroxamic acid peptide inhibitors showed differential effects. The thiols were equipotent inhibitors of both the collagen- and proteoglycan-degrading enzymes whereas the carboxyalkyls appear to inhibit solely the proteoglycan-degrading enzyme(s). The hydroxamic acid peptides, the most potent inhibitors, appear to be more active against the proteoglycan-degrading enzymes. These synthetic inhibitors of proteoglycan- and/or collagen-degrading enzymes may represent a new class of disease-modifying antiarthritic agents.

Animals↗
PubMed1986

Lack of response of rat chondrosarcoma chondrocytes to interleukin-1 to produce proteoglycan- and collagen-degrading enzymes.

Chondrocytes obtained from rat articular cartilage and Swarm rat chondrosarcoma were maintained in culture. Human interleukin-1 (IL-1) was added at confluency, and three days later the spent media was evaluated for proteoglycan- and collagen-degrading enzymes. Unlike chondrocytes from articular cartilage from several species, chondrocytes from Swarm rat chondrosarcoma did not respond to IL-1 stimulation. This lack of sensitivity to IL-1 may be a useful biochemical marker to classify various types of chondrosarcomas. However, it appears that the use of chondrosarcoma chondrocyte cultures are not useful as a source for metalloproteinases.

Animals↗
PubMed1985

Effect of antirheumatic agents on the mitogen response of arthritic rat spleen cells.

Splenic lymphoid cells from rats with adjuvant induced polyarthritis show a diminished response to the T-cell mitogens Con A and PHA. This report describes the in vitro effects of various antirheumatic agents on the mitogen induced proliferative response of normal (NSC) and arthritic (ASC) rat spleen cells. Indomethacin enhanced only the arthritic responses. Other PG synthesis inhibitors such as ibuprofen and naproxen enhanced the arthritic as well as the normal spleen cell proliferative responses. Gold sodium thiomalate augmented arthritic but not normal blastogenesis. Penicillamine significantly enhanced only the Con A response of arthritic cells at 10(7)M. Levamisole produced a significant increase in the PHA response of arthritic cells and the Con A NSC response. Chloroquine diphosphate enhanced the Con A response of normal cells at 10(-5)M and 10(-6)M; both chloroquine and tilorone suppressed blastogenesis of arthritic spleen cells at 10(-4)M and 10(-5)M. When classifying antirheumatic agents as stimulator or suppressors of immune function, one must account for their effects on arthritic as well as normal lymphoid cells at the predicted pharmacologic plasma levels.

Animals↗
PubMed1985

Comparison of spontaneous and retinoic acid stimulated rabbit articular cartilage degradation in vitro.

The in vitro degradation of rabbit articular cartilage explants was evaluated with and without the addition of retinoic acid under various experimental conditions. Retinoic acid at nontoxic concentrations ranging from 1 X 10(-7) to 1 X 10(-5) M significantly increased cartilage degradation. The addition of phenanthroline or cycloheximide, but not pepstatin, significantly inhibited spontaneous and retinoic-acid-stimulated cartilage degradation at pH 7. When the pH was reduced to 5, only pepstatin inhibited spontaneous and retinoic-acid-stimulated cartilage degradation. No chondroitin sulphate release was observed when the temperature was reduced to 4 degrees C. The different inhibitory profiles observed at pH 7 and pH 5 suggest that cartilage degradation at pH 7 is associated with the presence and synthesis of a neutral metalloproteinase.

Animals↗
PubMed1977

Inhibition of arachidonic acid induced mortality in rabbits with several non-steroidal anti-inflammatory agents.

The intravenous administration of arachidonic acid to rabbits is an effective in vivo model for evaluating potential anti-thrombotic drugs. Most of the non-steroidal anti-inflammatory agents (NSAIFA) inhibit this arachidonic acid induced mortality (except sodium salicylate and acetaminophen). However, there is a lack of correlation between the relative potencies from various assays (rabbit anti-thrombotic, anti-inflammatory, alalgesic, ulcerogenic and inhibition of prostaglandin synthetase evaluations). These studies imply other actions with NSAIFA than an effect solely on the prostaglandin biosynthetic pathway.

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