Proteolytic mechanisms operating at the surface of invasive cells.
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Biomedical subjects
Publications and source records attributed to R Stephens.
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A new screening test is described which enabled rapid determination of the proportion of single-chain and two-chain urokinase produced in the culture supernatants of 18 human cell lines. A clear distinction was found between two groups of cell lines: cells derived from ten solid tumors produced almost exclusively single-chain proenzyme, while the majority of the enzyme found in cultures of eight leukemia cell lines was in the active, two-chain form.
A retrospective survey of 612 eyes that had undergone cataract extraction and IOL implantation was undertaken to evaluate the accuracy of ultrasound biometry combined with keratometry using the SRK regression formula, for the preoperative prediction of intraocular lens powers. A mean error of +0.35 dioptre sphere (DS) (SD +/- 0.98) was found for the series overall, with a significant (P less than 0.005) difference between the distribution of postoperative refractive errors using the S.R.K. formula for IOL prediction and the use of a standard lens of 19.5 DS. The consistency of results was tested for those patients with greater or less than normal axial length. Linear regression analysis showed no correlation between axial length and postoperative refractive error and therefore does not support the adjustment of predicted IOL powers by a factor based on axial length. Statistically significant differences were found between surgeons' results, supporting the practice of A-constant modification for individual surgeons.
Specific allergens were applied topically to the conjunctivae of 18 allergic patients. In cases with positive allergic reaction, proteolytic activity, identified as plasmin using zymographic analysis and a monoclonal anticatalytic antibody to plasmin, was found to appear within 3-5 min in the tear fluid, reaching concentrations of 2.2-28.6 micrograms/ml. No plasmin was detected, either in the tear fluid of 30 unchallenged non-atopic control persons, or atopic patients provoked with a test material that had given no reaction in a skin prick test. These findings suggest that generation of plasmin is a consequence of the conjunctival allergic reaction.
We report a convenient sensitive enzyme activity assay for urokinase and tissue-type plasminogen activators, based on a solid-phase microtitre plate method using readily available polyclonal antibodies. The sensitivities for urokinase (active and proenzyme) and tissue activator were better than 1 ng/ml. The specificity was very high, with no significant contribution of urokinase in tissue activator assays or vice versa. This method is particularly useful for the assay of urokinase proenzyme in samples containing inhibitors. We describe how this assay may also be used to measure specific inhibitors of plasminogen activators, making use of their rapid formation of stable complexes with solid-phase activator. Inhibitors may be assayed in samples containing proenzymes.
Neutrophil function was assessed in patients undergoing anesthesia and surgery using a chemiluminescence (CL) assay. With the anesthetic agents enflurane and nitrous oxide, peroperative CL (99.1 mV; 13.8 SEM: postinduction but prior to surgery) was significantly lower than the preoperative value (146.5 mV; 14.1 SEM) with a mean fall of 30% (P less than 0.001). CL measurements taken 24 hr postoperatively were significantly increased (193.9 mV; 16.4 SEM) over the pre- and peroperative values, showing mean increases of 32 and 96%, respectively (P less than 0.001 in both cases). The inhibitory influence on CL appeared to be due to serum factors since peroperative patients' sera inhibited control neutrophils. Significantly depressed levels of the complement component C3 and IgG detected during the peroperative period (P less than 0.05) may explain this phenomenon. Postoperatively, C3 and IgG levels returned to normal. The transient decrease in peroperative neutrophil function may be a contributory factor to the establishment of postoperative sepsis in surgical patients.
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The prune belly syndrome is a congenital set of anomalies that includes cryptorchidism. Despite the known risk of testicular tumors in cryptorchid testes, what may be the first case of a germ cell tumor complicating the prune belly syndrome is described herein.
Four hundred forty previously untreated patients with active multiple myeloma were entered into a randomized trial (Southwest Oncology Group [SWOG] study 7927/28) comparing vincristine, melphalan, Cytoxan (Mead Johnson & Company, Evansville, Ind), and prednisone (VMCP) alternating with vincristine, BCNU, Adriamycin (Adria Laboratories, Columbus, Ohio) and prednisone (VBAP) with or without levamisole with vincristine, Cytoxan, and prednisone (VCP) with or without levamisole for induction therapy. The treatment groups were well balanced for all of the known major prognostic factors. Patients receiving VMCP-VBAP responded (greater than or equal to 75% regression) more frequently to induction therapy, both without (54%) and with (44%) levamisole v VCP without (28%) or with (28%) levamisole (P less than .001). In addition, patients receiving VMCP-VBAP (+/- levamisole) had a survival duration determined to be significantly increased by all forms of analysis: 48 and 33 months for VMCP-VBAP without and with levamisole v 29 and 26 months for VCP without and with levamisole (P = .011 overall). Levamisole did not improve response rates or survival duration (P greater than or equal to .1), nor did it prolong remission in the maintenance phase (P = .85). Analysis of SWOG study 7704/05 (updated April 1985) confirmed improved survival for combination therapy v MP, but no benefit for levamisole. The overall findings support the use of VMCP-VBAP as an excellent treatment option for remission induction in patients with active myeloma of all stages and prognostic categories.
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Dispersed single-cell suspensions of human gastrinoma tissue were incubated for 15, 60, and 120 min in a calcium-containing medium, (0.1, 2, 10 mM) in calcium-free medium and in calcium-free medium containing the calcium ionophore A23187 (0.01, 1, and 100 micrograms/ml). Supernatant and pellet (intracellular) gastrin levels were determined by radioimmunoassay. Supernatant gastrin levels remained stable over 120 min in calcium chloride or calcium gluconate containing medium, while intracellular pellet gastrin approximately tripled during the same incubation period. Total gastrin (supernatant plus pellet) approximately doubled during the 2-hr incubation in calcium. However, calcium (0.1, 2, or 10 mM) failed to produce a dose-dependent rise in supernatant, pellet, or total gastrin when compared to calcium-free incubates. Contrary to the expected gastrin response to calcium, supernatant and pellet gastrin levels were higher in incubates in calcium-free medium than in calcium-containing incubates. A23187 (0.01 or 1 mcg/ml) in a calcium-free medium decrease supernatant gastrin while high dose ionophore (100 mcg/ml) increased supernatant gastrin. All doses of ionophore stimulated pellet and total gastrin levels. Thus, it appears that the clinical augmentation of gastrin levels, seen with calcium challenge in vivo may not be solely due to changes in serum calcium.
8-Methoxypsoralen (8-MOP) and 4,5',8-trimethylpsoralen (TMP) were assessed for their effects on cyclic AMP (cAMP) accumulation in normal human fibroblasts, epithelial cells, and mononuclear leukocytes in vitro. In the absence of ultraviolet A radiation, 8-MOP and TMP caused an increase in the concentration of cAMP, but not cyclic GMP, in all of these cells. The effect was seen within 1 min and was sustained over control levels for at least 6 h. At 15 min, the highest dose of 8-MOP or TMP increased the cAMP concentration of fibroblasts by 52% and 69%, respectively. In mononuclear leukocytes, 8-MOP led to a 142% increase in cAMP after 20 min. Several analogs of 8-MOP and TMP had no effect on the cyclic nucleotide system of the cells. The pattern of the time-dependent increase in cAMP indicated that the effect was probably due to an inhibition of phosphodiesterase, rather than an activation of adenylyl cyclase. These results demonstrate that clinically effective psoralens, in the absence of UV radiation, can alter the cAMP concentration of human fibroblasts, epithelial cells, and mononuclear leukocytes in vitro.
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To simplify the diagnosis of chlamydial genital infection, we used a fluorescein-conjugated monoclonal antibody in immunofluorescence tests on smears prepared from urethral or cervical secretions obtained directly from patients. This direct test, requiring less than 30 minutes to perform, was based on the detection of extracellular chlamydial elementary bodies. A comparison of the direct test with cultures stained with iodine on specimens from 926 patients demonstrated a sensitivity of 93 per cent and a specificity of 96 per cent. The direct test provides a rapid, simple, and sensitive method for the diagnosis of chlamydial infection, which can be performed in laboratories that do not have tissue-culture capability.
Pentamidine can cause hypoglycemia followed by hyperglycemia. The mechanism of this biphasic response is not known but has been reported to be similar to that of streptozotocin and N-3-pyridyl-N-p- nitrourea (Vacor). Pentamidine (4 mg/kg per day for 12 days) was used in a patient with malignant insulinoma after several unsuccessful debulking procedures and chlorozotocin therapy. Mean glucose and immunoreactive insulin levels (+/- SEM) before and after therapy were 80 +/- 40 mg/dl versus 70 +/- 50 mg/dl and 216 +/- 12 microU/ml versus 198 +/- 22 microU/ml, respectively. These were not significantly different. The patient's five-month-old malignant insulinoma monolayer cell culture was incubated with pentamidine (60 micrograms/ml) in the presence or absence of supplemented stimulatory medium consisting of glucose (300 mg/dl) and theophylline (20 micrograms/ml). Chloroquine (60 micrograms/ml) was added to inhibit lysosomal degradation of immunoreactive insulin. Aliquots of media for immunoreactive insulin determination were obtained at 30 minutes, 20 hours, 72 hours, and three weeks. The cells were examined by high-power light microscopy at each time interval. At 30 minutes, pentamidine alone caused passive release of immunoreactive insulin, 23 percent higher than control (p less than 0.01). Stimulatory medium increased immunoreactive insulin 45 percent greater than control (p less than 0.01). Pentamidine plus stimulatory medium had no additive effect on immunoreactive insulin released within 30 minutes. At the end of 20 hours, immunoreactive insulin was no different with pentamidine and/or stimulatory medium. However, the addition of chloroquine increased immunoreactive insulin by 35 percent above the medium with pentamidine and stimulatory medium (p less than 0.01). At 72 hours, pentamidine suppressed immunoreactive insulin by 100 percent in all the media, irrespective of the presence or absence of stimulatory medium and/or chloroquine. At the end of three weeks, there was 50 percent suppression of immunoreactive insulin in the control medium, but pentamidine again completely suppressed immunoreactive insulin. High-power microscopy demonstrated intact cells in the control medium, whereas no cell structure could be detected in the media containing pentamidine at three weeks. In summary, pentamidine had no acute in vivo effect in a patient with malignant insulinoma. However, when used in an in vitro monolayer system, pentamidine caused (1) acute immunoreactive insulin release followed by inhibition of immunoreactive insulin secretion and (2) cytolysis of human malignant insulinoma cells in vitro.
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Twenty-six evaluable patients with advanced soft tissue and bony sarcomas refractory to chemotherapy were treated with vincristine plus high-dose methotrexate and leucovorin rescue. A 14% response rate was observed among 14 patients presenting with refractory soft tissue sarcomas. No responses were observed among 12 patients with bony sarcoma. Toxic reaction with nausea, vomiting, nephrotoxicity, and myelosuppression was manageable. While this study did demonstrate activity of this regimen in doxorubicin-refractory patients, the duration of the responses was relatively brief. Thus, the clinical utility of such a regimen is questionable.