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K Pritchard

Publications and source records attributed to K Pritchard.

47 records · Page 3Linked to original sources
PubMed1987

The mechanism of Ca2+ regulation of vascular smooth muscle thin filaments by caldesmon and calmodulin.

The interactions of vascular smooth muscle caldesmon with actin, tropomyosin, and calmodulin were determined under conditions in which the four proteins can form reconstituted Ca2+-sensitive smooth muscle thin filaments. Caldesmon bound to actin in a complex fashion with high affinity sites (K = 10(7) M-1) saturating at a stoichiometry of 1 per 28 actins, and lower affinity sites at 1 per 7 actins. The affinity of binding was increased in the presence of tropomyosin, and this could be attributed to a direct interaction between caldesmon and tropomyosin which was demonstrated using caldesmon cross-linked to Sepharose. In the presence of tropomyosin, occupancy of the high affinity sites was associated with inhibition of actin-activated myosin MgATPase activity. Caldesmon was found to bind to calmodulin in the presence of Ca2+, with an affinity of 10(6) M-1. The binding of Ca2+ X calmodulin to caldesmon was associated with the neutralization of inhibition of actin-tropomyosin. Ca2+ X calmodulin binding reduced but did not abolish the binding of caldesmon to actin-tropomyosin. From this data we have proposed a model for smooth muscle thin filaments in which Ca2+ regulates activity by converting the inhibited actin-tropomyosin-caldesmon complex to the active complexes, actin-tropomyosin-caldesmon-calmodulin X Ca2+ and actin-tropomyosin.

Actin Cytoskeleton↗
PubMed1987

Evidence for Na+/Ca2+ exchange in isolated smooth muscle cells: a fura-2 study.

Isolated smooth muscle cells (SMC) from guinea pig taenia coli were employed. Suspension of cells were externally loaded in saline with the fluorescent calcium indicators quin-2/AM or fura-2/AM at 20-40 microM or 4 microM respectively, resulting in an estimated intracellular concentration of 100-200 microM for quin-2 or 10-20 microM fura-2 (free acid). On addition of 100 microM carbachol or high K+o (80 mM) depolarization, fura-2 loaded cells contracted (104 +/- 47 micron, n = 121 rest: 39 +/- 13 micron, n = 59 contracted) identically to control (103 +/- 35 micron, n = 232 rest: 39 +/- 16 micron, n = 89 contracted) cells, whereas quin-2 loaded cells were unresponsive to these protocols and there was no significant length change. The Ca2+i of fura-2 loaded cells was 100 +/- 18 nM (mean +/- SD, n = 15) and was not significantly different from quin-2 loaded cells 107 +/- 26 nM (n = 13). Treatment of fura-2 loaded cells with 100 microM ouabain saline for 10-60 min progressively elevated the Ca2+i to a mean of 266 +/- 83 nM (n = 15). Reduction of Na+o (96% Li+ replaced) significantly increased Ca2+i to 317 +/- 77 nM (n = 8). After pretreatment with ouabain (100 microM), Na+o replacement (Li+) increased Ca2+i at a significantly faster rate [3.6 nM min-1 (control) cf. 19.8 nM min-1 (ouabain)].

Animals↗
PubMed1986

Na+/Ca2+ exchange in isolated smooth muscle cells demonstrated by the fluorescent calcium indicator fura-2.

Fura-2, a novel fluorescent indicator of cytoplasmic calcium concentrations ([Ca2+i]), was 'loaded' into smooth muscle cells isolated from guinea pig taenia coli. Resting cells maintained a stable [Ca2+i] of 107 +/- 26 nM (n = 13), which could be perturbed with ionomycin. [Ca2+i] was elevated by stimulation of the cells with carbachol or 50 mM KCl. Reduction of the plasmalemmal Na+ concentration gradient by inhibition of the Na+/K+-ATPase with ouabain markedly elevated [Ca2+i]; this elevation was dependent on extracellular Ca2+. [Ca2+i] was also increased by replacement of the extracellular Na+ with an organic cation.

Animals↗
PubMed1986

Caldesmon: a calmodulin-binding actin-regulatory protein.

The protein caldesmon, originally isolated from smooth muscle tissue where it is the most abundant calmodulin-binding protein, has since been shown to have a wide distribution in actin- and myosin- containing cells where it is localized in sub-cellular structures concerned with motility, shape changes and exo- or endo-cytosis. Caldesmon is believed to be an actin- regulatory protein, and binds with high affinity to actin or actin-tropomyosin. Caldesmon inhibits the activation by actin-tropomyosin of myosin MgATPase activity, and the inhibition can be reversed by Ca2+.calmodulin. The binding of caldesmon to smooth muscle proteins has been studied in detail, enabling a model to be constructed which could account for the observed Ca2+ regulation of smooth muscle thin filaments. The abundance of caldesmon, and the Ca2+-regulation of its activity via calmodulin, mean that it is potentially an important intracellular regulator of processes such as smooth muscle contraction, cell motility and secretion.

Actins↗
PubMed1986

Phase II study of lonidamine in patients with metastatic breast cancer: a National Cancer Institute of Canada Clinical Trials Group Study.

The National Cancer Institute of Canada Clinical Trials Group conducted a phase II study of lonidamine, given in an escalating oral daily schedule to a maximum dose of 450 mg/m2 in patients with previously treated advanced breast cancer. Five responses were seen in 30 evaluable patients (17%). Treatment was discontinued because of toxicity in seven patients. Toxicity generally consisted of myalgia, nausea, vomiting, skin hyperesthesia, somnolence, and ototoxicity. All side effects were reversible and no hematologic toxicity was observed. The absence of myelosuppression and the suggestive lack of cross-resistance between lonidamine and standard chemotherapeutic drugs warrant further studies of lonidamine in breast cancer, particularly in combination with other agents.

Adult↗
PubMed1985

A phase II study of spirogermanium in patients with metastatic malignant melanoma. An NCI Canada Clinical Trials Group Study.

The National Cancer Institute of Canada Clinical Trials Group conducted a phase II study of spirogermanium given daily for 5 days every 3 weeks to previously untreated patients with malignant melanoma. In 21 evaluable patients one complete response was seen (response rate 5%). Disease progression occurred in the other 20 patients. Toxicity was primarily neurologic and mild or moderate in most patients, though there was one treatment related death. In this schedule spirogermanium has extremely limited activity against malignant melanoma and will not contribute significantly to the systemic therapy of this disease.

Adult↗