Proteinase and proteinase-inhibitor activities of rat uterine myometrium during pregnancy and involution

Abstract

A supernatant fraction was prepared from rat uterine myometrium by homogenization, sonication and centrifugation. In this supernatant the protein concentration and the activities of an acid proteinase, an acid phosphatase and a proteinase inhibitor were measured. From the fibrous sediment, after washing with 0.5% Triton X-100 and with water, an actomyosin-containing solution was obtained by extraction with 0.6M-NaCl, and in this extract the protein concentration and a neutral proteinase activity were measured. The myometrial wet weight and the activities of the acid proteinase, acid phosphatase and proteinase inhibitor increased by factors of 3-15 during pregnancy and decreased to the same or a greater extent during involution. The amount of protein extracted with 0.6M-NaCl increased by a factor of only 2.3 and the neutral proteinase activity remained essentially constant during pregnancy and involution. The pH optimum of the neutral proteinase, and its pattern of activity compared with those of the lysosomal enzymes, show that the neutral proteinase is not of lysosomal origin. Actomyosin is degraded by the neutral proteinase activity in vitro. Since actomyosin is rapidly broken down only after parturition, the action of the neutral proteinase activity on actomyosin, if this occurs in vivo, must be regulated in some way. The proteinase-inhibitor activity measured in the first supernatant varied in a manner which suggested that it could be involved in this control.

Oestrogenic activity of tamoxifen and its metabolites on gene regulation and cell proliferation in MCF-7 breast cancer cells

“…Although the biological role of cathepsin D in breast tumours is not known, it has been proposed to be involved in tumour dissemination as it can be secreted from breast tumour cells (Westley & Rochefort, 1980) and can degrade extracellular matrix (Briozzo et al, 1988). The present observations suggest that tamoxifen treatment might enhance any tumour dissemination mediated by cathepsin D. However, cathepsin D is also thought to be involved in tissue involution and remodelling; and in processes such as post-partum involution of the uterus, cathepsin D levels are under some degree of hormonal control (Afting et al, 1979). Cathepsin D may therefore play an active role in tumour regression and the dramatic elevation of cathepsin D by tamoxifen in the presence of oestradiol might facilitate this process.…”

Production of Angiotensin II by Homogeneous Cultures of Vascular Smooth Muscle Cells From Spontaneously Hypertensive Rats

“…The cells were centrifuged for 10 minutes at 12 000g and resuspended in 1 mL of 50 mmol/L Tris-HCl (pH 7.4) containing 10 mmol/L mercaptoethanol. After sonication for four 15-s bursts, the protein concentration of the cell lysate was measured by the method of Lowry et al 35 The lysate was centrifuged for 10 minutes at 12 000g, and the resulting supernatant was assayed for acid protease activity as described by Afting et al 37 Briefly, 60 mL of the cell extracts in the absence or presence of 1 mmol/L pepstatin A (an inhibitor of aspartyl proteases) was incubated at 37°C for 30 minutes, with 80 mL of borate buffer (50 mmol/L Na 2 B 4 O 7 [pH 8.6] and 1 mol/L NaCl) containing hemoglobin (10 mg/mL) as substrate. The reaction was stopped by the addition of 100 mL of 10% (wt/vol) trichloroacetic acid.…”

Cytosol cathepsin-D content and proliferative activity of human breast cancer

“…This suggests that high levels of cath-D could indicate a favorable prognosis for breast cancer patients. To support this hypothesis, we must recall that cath-D is thought to be involved in the physiological phenomena of tissue involution and remodelling [21], such as postpartum uterine modifications. Furthermore, cath-D might play a direct role in tumor regression, as demonstrated by the evaluation ofmRNA and protein cath-D levels in tamoxifen treated cell lines [22] and breast cancer patients [23].…”