[Reconstruction of the extensor apparatus with advanced structural defects in knee revision arthroplasty]. / Rekonstruktion des Streckapparates bei fortgeschrittenen Defekten in der Knierevisionsendoprothetik.

OBJECTIVE: Restoration of the extensor apparatus in the case of advanced tendon defects as part of revision total knee arthroplasty (TKA). Reconstruction and augmentation using vascularized gastrocnemius muscle and tendon. INDICATIONS: Advanced degeneration of the extensor apparatus (patella tendon; quadriceps tendon) with or without discontinuity, following revision arthroplasty. CONTRAINDICATIONS: Persistent infection or pending TKA revision. Damaged gastrocnemius or soleus muscle or Achilles tendon. SURGICAL TECHNIQUE: Extension of the surgical TKA-access medial-distally. Separation of the medial gastrocnemius muscle along the raphe and preparation of the distal tendon from the soleus portion. Transposition into the defect site, augmentation or reconstruction of the defect by double turn of the gastrocnemius tendon. The muscle belly serves to adequately cover the tendon as well as the ventral knee joint. Mesh coverage of the muscle. POSTOPERATIVE MANAGEMENT: Immobilization of the knee and ankle for 10 days until mesh graft healing. Stepwise increasing flection of the knee with 30°/60°/90° every 2 weeks. Total weight bearing with secured full extended knee, no weight bearing with flexed knee for 6 weeks. RESULTS: In 9 patients, 3 with complete rupture of the patellar tendon, 5 with destruction of the extensor apparatus, and 1 patient with rupture of the quadriceps tendon following TKA revision, good functional results were achieved with active extension of the knee joint and standing/gait stability 6 months after surgery.

[Modified percutaneous achillotentomy according to Hoke for minimally invasive treatment of pointed foot]. / Modifizierte perkutane Achillotenotomie nach Hoke zur minimal-invasiven Spitzfußbehandlung.

OBJECTIVE: Various underlying diseases can lead to a pointed foot in children and adults. If the gastrocnemius and soleus muscles are structurally shortened, various surgical procedures are available to correct this malposition. A preferred method for restoring a normal dorsiflexion of the upper ankle joint is percutaneous achillotentomy according to Hoke. Consideration of the physiological-anatomical torsion of the Achilles tendon as it corresponds to the White technique and is recommended by some authors shows in our experience no advantages. In the present work, we show a modified, likewise minimally invasive form of this surgical method with which immediate full weight-bearing of the affected lower extremity is possible from postoperative day 1. INDICATIONS: All clinically relevant structural pointed foot, for primary and/or for revision treatment. CONTRAINDICATIONS: Infection in the area of the operation. SURGICAL TECHNIQUE: No tourniquet, 3 incisions with the 15 mm knife: (1) medial distal at the transition from the Achilles tendon to the calcaneus, (2) medial proximal approximately 7 cm proximal to the 1st stab incision, (3) lateral, midway between the first two incisions; no skin suturing, application of a lower leg cast. POSTOPERATIVE MANAGEMENT: On postoperative day 1, cast hybridization using Scotchcast (3M Deutschland GmbH, Neuss, Germany), followed by pain-adapted full weight-bearing; removal of the cast in the outpatient department after 4 weeks. RESULTS: A total of 104 patients underwent surgery, 1 case of a local pressure point, no infections, no overcorrections, no Achilles tendon ruptures, in one case a postoperative relapse due to a broken cast. The risk of overcorrection to the foot, which was considered the main complication in the literature, did not occur in any of the cases.

Endogenous and synthetic steroids in bovine urine--preparation of in-house reference material, stability studies and results of a proficiency test.

Within the framework of the German National Residue Control Plan a specific number of samples of animal origin have to be analysed for natural and synthetic steroids each year. As a measure of external quality control of the methods applied in routine analysis a proficiency test was carried out. To this end, in-house reference material containing incurred residues of 17α- and 17ß-nortestosterone and 17α- and 17ß-estradiol as well as fortified residues of 17α-methyltestosterone and 17α-trenbolone in bovine urine were produced. Before sending the proficiency test material to the participants, the homogeneity of all samples was tested and confirmed. Furthermore extensive short- and long-term stability studies were carried out. The statistical evaluation of the proficiency test was performed by applying robust statistics as described in standard DIN 38402. Based on the target value and standard deviation z-scores were calculated as standardised measure of the laboratory performance. The evaluation of the proficiency test showed that nine laboratories submitted quantitative results within the tolerance limits for all analytes. Taking into account the individual decision limits, there were no false negative results. In overall evaluation, 11 of 12 laboratories participated successfully.

Endosonographic diagnosis of solid pancreatic tumors: a retrospective analysis from a tertiary referral center.

BACKGROUND: The sensitivity of endosonography for pancreatic tumors is good, but differentiation between malignant and benign lesions remains a challenge. We, therefore, analyzed the correlation between endosonographic findings and pancreatic carcinoma in a population with a high prevalence of chronic pancreatitis. METHODS: 171 endosonographic examinations were retrospectively evaluated using 22 dichotomous criteria. Final diagnosis was defined by the results of pancreas resection or by clinical follow-up (median: 41 months). A sum score was established after multivariate analysis. RESULTS: Final diagnosis was carcinoma, chronic pancreatitis, neuroendocrine tumor and normal pancreas in 67, 65, 9, and 38 subjects (prevalence 39 %, 38 %, 5 %, 22 % respectively) After multivariate analysis, carcinoma was significantly correlated with six endosonographic criteria and age, which resulted in the following score (yes = 1, no = 0): (dilated pancreatic duct) + (vascular infiltration) + (suspicious lymph nodes) + (tumor edge with pseudopodia-like extensions) + (age > 50 years) - (increased pancreas parenchyma echogenicity) - (tumor diameter < 10 mm) + 3. After receiver operating characteristic analysis, the area under the curve was 0.85. For score values of 5 (4) or more, sensitivity was 0.63 (0.93), specificity 0.91 (0.55), positive predictive value 0.82 (0.57), negative predictive value 0.79 (0.92), positive likelihood ratio 7.24 (2.05), and negative likelihood ratio 0.41 (0.14). CONCLUSION: A simple and potentially useful score for the prediction of pancreatic cancer based on six endosonographic criteria and patient age was established. Distribution of final diagnoses in the patient population argues for the score's applicability in clinical practice of a tertiary referral center and warrants prospective validation.

Determination of chloramphenicol residues in crustaceans: preparation and evaluation of a proficiency test in Germany.

Chloramphenicol (CAP) is banned for use in food-producing animals and is, thus, controlled on the basis of the National Residue Control Plans in the European Union. Due to current problems with residues of CAP in shrimp, crayfish and prawns, a sensitive GC/NCI/MS method was optimised and in-house validated. The validation study resulted in a decision limit (CCalpha) of 0.07 microg kg-1, a recovery of 95% and a within-laboratory reproducibility of 9%. The method was used for preparing a proficiency test to assess the quality of residue control in Germany. The proficiency test involved analysis of five samples and the results were very satisfactory. The reproducibility standard deviation for five samples ranged from 17 to 24%, and the median concentrations lay between 0.43 and 0.51 microg kg-1 CAP. These values are clearly below the corresponding Horwitz standard deviation of about 50%. From the study, it can be concluded that there are, irrespective of the method applied, well-established and proper working analytical procedures for the control of CAP around the minimum required performance limit (MRPL) of 0.3 microg kg-1.

Introital ultrasound of the lower genital tract before and after colposuspension: a 4-year objective follow-up.

OBJECTIVE: To assess the topography of the bladder neck by introital ultrasound before and after open colposuspension. METHODS: Three hundred and ten women with urodynamically proven stress urinary incontinence were included in this long-term study to investigate the position and function of the bladder neck at rest and during straining. Height (H), distance (D), and urethrovesical angle of the bladder neck (beta) were measured by means of preoperative and postoperative introital ultrasound. Women were followed up; 152 of them (49%) completed 48 months of follow-up. RESULTS: At the 6-month follow-up examination, 90.0% of the women were continent (279/310), 3.5% (11/310) showed voiding difficulties, 3.5% (11/310) had urgency, and 1.6% (5/310) had developed de novo urge incontinence. At the 48-month follow-up, 76.8% of the patients were still continent. All postoperative measurements yielded significantly lower values for angle beta at rest and during straining compared with the preoperative results (P < 0.0001). The median linear movement of the bladder neck during straining decreased from 18.0 mm before surgery to 6.4 mm at the 48-month follow-up (P < 0.0001). The median level of ventrocranial elevation of the vesicourethral junction was 14.3 mm immediately after surgery, 9.9 mm after 6 months and 6.6 mm after 48 months. The degree of surgical bladder-neck elevation was associated with postoperative urgency/de novo urge incontinence (P < 0.0001) and voiding difficulty (P < 0.0001). CONCLUSIONS: The colposuspension procedure reduces angle beta at rest and during straining, restricts linear movement with straining, and elevates the bladder neck. Perioperative introital ultrasound improves understanding of this surgical procedure and might help to prevent postoperative complications.

[Ultrasound imaging of the lower urinary tract in women before and after colposuspension: a 6-month follow-up]. / Introitussonographie des unteren Genitaltraktes vor und nach Kolposuspension: eine sechsmonatige Nachbeobachtung.

OBJECTIVE: To evaluate the position of the bladder neck before and after open Burch colposuspension, using introital ultrasound. DESIGN: Retrospective longitudinal study using pre- and post-operative sonographic assessment of the position and function of the bladder neck. SETTING: Urogynecology units at the universities of Marburg/Göttingen and Witten/Herdecke and the DRK district hospital in Alzey, Germany. PATIENTS: 310 women undergoing open Burch colposuspension for primary genuine stress incontinence between September 1992 and December 2001. METHOD: Two-dimensional introitus sonography of the bladder neck prior to, one week and six months after surgery. RESULTS: The median age at surgery was 55 years (26 - 85). Open colposuspension lead to a 90.0 % (279/310) cure rate at 6 months with only 3.5 % (11/310) of the patients showing persistent micturation problems. A further 11.6 % (36/310) had symptoms of urgency and in 7 patients (2.3 %) a de novo urge-incontinence occurred. Post-operative bladder neck angles and movements at rest and during valsalva manoeuvre were significantly reduced while the resting bladder neck position was significantly elevated (p < 0.0001). Anatomical elevation of the bladder neck after open colposuspension varied between 2 - 39 with a median of 14.3 mm of neck elevation after one week and 9.9 mm at 6 month, respectively. Incontinence surgery lead to a significant reduction of the urethral funneling (p < 0.0001). CONCLUSION: In our series, open Burch colposuspension decreased both the bladder neck angle and the linear movement at rest and on valsalva as a result of the surgically stabilized bladder neck. Thus, our results support the hammock hypothesis that even small changes in the position of neck position are sufficient to reverse incontinence. We believe that perioperative introitus sonography is a helpful tool for the clinical assessment and documentation of not only morphological but also functional changes of the female continence organ before and after open Burch colposuspension.

A comparison of EDS microanalysis in FIB-prepared and electropolished TEM thin foils.

This paper reports the results of a fine-probe EDS microanalytical study of cellular precipitation in a Cu-Ti binary alloy. Compositional profiles across the solute depleted Cu-rich FCC lamellae and the Cu4Ti lamellae within isothermally formed cellular colonies were measured in a FEG-TEM from thin-foil specimens prepared by conventional electropolishing and by a technique using a Ga+ focused ion-beam (FIB). The Cliff-Lorimer ratio method, with an absorption correction, was employed to quantify the compositions. Two FIB samples were prepared with different orientations of the lamellae with respect to the ion-milling direction. The compositional profiles across the Cu-rich FCC lamellae and the Cu4Ti compound lamellae in both the FIB-prepared samples and the electropolished sample were, within experimental error, numerically equivalent. The composition of the Cu4Ti compound phase lamellae was very close to the ideal stoichiometric composition of 20 at % Ti. It is concluded that for this system, and for the specimen preparation procedures used in this study, the Ga+ ion-milling process has had no detectable effect on the chemistry changes across the interlamellar interface at the scale studied. These results indicate that the possible sources of chemical artifacts which include redeposition, preferential sputtering and ion-induced atomic migration can be minimized if several precautions are taken during milling in the FIB. Consistent with previous investigators, it was also found that the ion-milling process does introduce significant structural artifacts (e.g., dislocations) into the softer FCC Cu-rich phase compared with a specimen produced by conventional electropolishing.

Regulation of GCDFP-15 expression in human mammary cancer cells.

Gross cystic disease fluid protein 15 (GCDFP-15) is a major protein component of benign breast gross cysts. It is also found in approximately 50% of all breast cancer specimens. Androgen receptor (AR) mediated regulation of GCDFP-15 expression was investigated in the AR-positive human mammary cancer cell lines MFM-223 and ZR-75-1. Proliferation of MFM-223 and ZR-75-1 cells is inhibited by androgens. Ten nM 5alpha-dihydrotestosterone stimulated the expression of GCDFP-15 mRNA in MFM-223 (ca. 3-fold) and ZR-75-1 cancer cells (ca. 30-fold) as well as the secretion of GCDFP-15 into the culture medium. Competition experiments with DHT and the antiandrogens hydroxyflutamide and casodex confirmed the involvement of the AR in the regulation of GCDFP-15. Both antiandrogens inhibited GCDFP-15 mRNA expression even in the absence of DHT. AR mRNA was down-regulated in MFM-223 and ZR-75-1 cells (80 and 20% of the control, respectively) during incubation with DHT. Our data demonstrate the effective inhibition of GCDFP-15 expression by pure antiandrogens.

Expression of placental protein 14 by the new endometrial cancer cell line MFE-280 in vitro and by endometrial carcinomas in vivo.

MFE-280 endometrial cancer cells express PP14 (placental protein 14) in vitro. PP14 is normally found in the secretory endometrium and in placental tissue. MFE-280 cells, which are tumorigenic in nude mice, were derived from a recurrent, poorly differentiated endometrial carcinoma. The cells were initially grown in suspension culture and later transferred to monolayer cultures. Karyotyping revealed near-diploidy with a complex heterogeneous aberration pattern. MFE-280 cells were positive for the cytokeratins 7, 8, 18 and 19 as well as for vimentin. The expression of PP14 in MFE-280 cells was demonstrated by immunochemistry and reverse transcriptase--polymerase chain reaction. PP14-mRNA was also detected in one out of five endometrial cancer specimen. In tumor tissue the expression of PP14 was not dependent on progestins.

Establishment of new epithelial carcinoma cell lines by blocking monolayer formation.

Human endometrial carcinoma cell lines were established by using a new cell culture technique. The malignant endometrial tumors were grossly disaggregated by mechanical means and cultivated in suspension culture. Adhesion to the bottom of the culture flasks was prevented by first coating the flasks with a thin agarose layer. Four cell lines were derived from 17 samples by this new technique. The cell lines obtained in this way were fully characterized, including karyotyping, intermediate filament staining and transplantation to nude mice. This new technique of initial suspension culture may also be applicable to other human tumors that are equally difficult to cultivate in vitro.

[Clinical and chemical laboratory course of HELLP syndrome--a retrospective analysis]. / Klinischer und laborchemischer Verlauf des HELLP-Syndroms--eine retrospektive Analyse.

The HELLP-Syndrom (hemolysis, elevated liver enzymes, low platelet count) is considered as a severe complication of eclampsia with unpredictable development of pregnancy including high maternal and fetal risk. The result of retrospective analysis of all deliveries of the years 1986-1991 at the UFK Marburg were 28 cases of proved HELLP-Syndrom. Medical history, correlation of clinical and laboratory findings as well as the development of the disease and the neonatal dates were evaluated by computerized documentation. The incidence of HELLP-Syndrom was 28 of 8111 deliveries at all (0,34%). 82% of the women with HELLP-Syndrom were primiparae. The leading symptom was right upper abdominal pain in 75%, which lasted already 5,7 days before presentation in the clinic. Hypertonus, edema and proteinuria were present in 71%, 61% and 89% of the cases. The diagnosis indicating laboratory finding was the thrombocytopenia (mean 62 G/l). In comparison to the thrombocytes, which were at the 4.-7. day pp in 89% within the normal range, the liver function tests normalized just between the 9. and 13. day pp (SGOT 89%, SGPT 77%). The shortening of the prepartal hospitalization from 6 days in 1986/87 to 8 hours in 1990/91 decreases the periand postnatal complication rate from 43% to 23%. 26/28 patients (92%) were delivered by caesarean section from healthy babies through which were 75% premature infants and in 27% of the cases small for gestational age additionally. We conclude that the decrease of the diagnosis-delivery interval and the intensive medical care are responsible for the diminution of the maternal and neonatal mortality rate to 0%.

Androgen-like and anti-androgen-like effects of antiprogestins in human mammary cancer cells.

In addition to their antiprogestational activity, the antiprogestins RU486, ZK98.299 and ZK98.734 possess varying antiglucocorticoid as well as androgen-like or antiandrogen-like properties in human mammary cancer cells. The human mammary cancer cell line MFM-223, which contains only androgen receptors, was used as a model to investigate androgen receptor mediated effects of these antiprogestins. Proliferation of MFM-223 cells is inhibited by androgens and does not respond to oestrogens, progestins and glucocorticoids. As shown in proliferation assays, ZK98.734 was a strong inhibitor of cell proliferation. This effect was antagonised by the antiandrogen hydroxyflutamide. ZK98.734 was found to displace [3H]R1881 from the androgen receptor in MFM-223 cells, substantiating the involvement of the androgen receptor. The antiprogestin ZK98.299 failed to influence the proliferation of MFM-223 cells. ZK98.299 did not bind to the androgen receptor and was devoid of androgenic or antiandrogenic activity. RU486 bound to the androgen receptor. It was a weak inhibitor of MFM-223 cell proliferation, but the inhibition of proliferation by RU486 was not antagonised by hydroxyflutamide. This effect was probably not mediated by the androgen receptor. RU486 had antiandrogenic activity in this cell line, as it antagonised the inhibitory effect of dihydrotestosterone at a 100-molar excess. These results were confirmed by transfection experiments with an MMTV-CAT construct in the same cell line, demonstrating the biological function of the ZK98.734-androgen receptor complex. ZK98.299 and RU486 were not able to induce CAT activity. The different androgenic or antiandrogenic properties of the antiprogestins investigated should be considered when selecting antiprogestational properties of the antiprogestins investigated should be considered when selecting antiprogestational compounds for clinical applications, as a partial androgenic activity may be of benefit in breast cancer but can have undesired side-effects in other diseases.

Androgen receptor mediated growth control of breast cancer and endometrial cancer modulated by antiandrogen- and androgen-like steroids.

Androgens are involved in many regulatory processes in mammary and endometrial epithelium, but their role in the development and progression of breast and endometrial carcinoma is poorly understood. Androgen receptors (AR) are found in normal epithelium as well as in more than 50% of specimen from both tumor types. The occurrence of AR is correlated with estrogen and progesterone receptors. Androgen receptor positive cell lines were established during the last few years in our laboratory from malignant mammary (MFM-223) and endometrial (MFE-296) tumors supplementing the small number of androgen-responsive cell lines published so far. In this paper some aspects of the role of androgens in these two types of hormone responsive female cancers are presented. The proliferation of ZR-75-1, MFM-223 and MFE-296 cells is inhibited by androgens. The progestin medroxyprogesterone acetate inhibits the proliferation of estrogen- and progesterone receptor negative MFM-223 cells via the androgen receptor. Some steroid metabolites with distinct estrogenic properties like androst-5-ene-3 beta,17 beta-diol possess androgenic properties in this model system. Androgens stimulate the in vitro secretion of gross cystic disease fluid proteins by human mammary cancer cells. These proteins are normally found in benign breast cysts in vivo. The occurrence of gross cystic disease is correlated with an increased risk of breast cancer. The AR is autoregulated in MFM-223 mammary cancer cells on the protein and mRNA level. In MFE-296 cells with endometrial origin AR protein was increased after incubation with androgens.

Androgen responsiveness of the new human endometrial cancer cell line MFE-296.

MFE-296 endometrial cancer cells express androgen receptors in vitro. These cells, which are tumorigenic in nude mice, are derived from a moderately differentiated human endometrial adenocarcinoma. They express vimentin and the cytokeratins 7, 8, 18, and 19. Karyotyping revealed near-tetraploidy for most of the cells. No marker chromosomes were observed. DNA analyses confirmed the genetic identity of the cell line and the patient from whom the cell line was derived. Proliferation of MFE-296 cells was inhibited by the progestin R5020 and the androgen dihydrotestosterone (DHT). The inhibition of proliferation by DHT was antagonized by the antiandrogen Casodex, demonstrating the involvement of the androgen receptor. Androgen binding was determined at 22,000 binding sites per cell using a whole-cell assay (KD = 0.05 nM) and 30 fmol/mg protein with the dextran charcoal method; 7 fmol/mg protein of progesterone receptors were found, whereas estrogen receptors were below 5 fmol/mg protein. The androgen receptor was functionally intact, as demonstrated by transfection experiments with a reporter-gene construct, containing an androgen-responsive element. In MFE-296 cells the content of the androgen receptor was up-regulated by its own ligand.

Production of insulin-like growth factor binding proteins by human ovarian carcinoma cells.

Cells of the human ovarian carcinoma lines EFO-21, EFO-27, MFO-35 and MFO-36 secrete binding proteins for insulin-like growth factors (IGFBPs) into their culture media. By sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS-PAGE) and ligand blotting, seven groups of IGFBPs with molecular masses of 25, 30 (doublet), 34, 37, 40, 45, and 50 kDa were observed, depending on the cell line under investigation. By Northern blot analyses using cDNAs or oligonucleotides specific for the six types of IGFBP (IGFBP-1 to IGFBP-6), mRNA for all IGFBPs tested except for IGFBP-1 could be detected in the ovarian carcinoma cell extracts. In detail, analysis of EFO-21 protein products by SDS-PAGE yielded IGFBPs of 25, 34, and 50 kDa; extracts of EFO-21 cells contained mRNAs for IGFBP-2, -3, -4, and -6. EFO-27 cells produced IGFBPs of 40 kDa and 45 kDa as determined by SDS-PAGE, and mRNAs for IGFBP-3, -4, and -6 were detected. In the conditioned medium of MFO-35 cells, IGFBPs of 25, 30 (doublet), 34, 37, 40, and 45 kDa were observed by SDS-PAGE, while mRNAs for the five proteins IGFBP-2 to IGFBP-6 were found. MFO-36 cells produced IGFBPs of 34 kDa and 50 kDa as determined by SDS-PAGE, and the cells expressed mRNAs for IGFBP-2, -3, -4, and -6. In relation to published molecular mass data of the known IGFBPs, the size of the secreted proteins could be correlated to the mRNA patterns expressed by the ovarian carcinoma cells. It is concluded that ovarian carcinoma cells frequently express IGFBP-3, -4, and -6 and, to a lesser extent, IGFBP-2; the expression of IGFBP-5 appears as a rather rare event, while IGFBP-1 was not found to be expressed in ovarian carcinoma cells.

Urinary excretion of growth factors in patients with ovarian cancer.

The levels of epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha) were analysed in 24-h urine samples from patients with ovarian malignancies, benign ovarian tumours, and healthy controls by specific radioimmunoassays. No significant difference in total urinary immunoreactive EGF excretion between the groups was detected. However, 79% (23/29) of the patients with ovarian carcinomas excreted TGF-alpha (median 12.6 pmol/24 h), whereas only 17% (2/12) of the patients with benign ovarian tumours and 23% (3/13) of the controls did so. The difference between cancer patients and controls was highly significant (P < 0.001). Analyses of the urine samples separated by gel filtration revealed a greater molecular heterogeneity of EGF and TGF-alpha in cancer patients than in controls. High and low molecular weight forms of EGF were able to bind to the EGF receptor and to induce anchorage-independent growth. After surgical reduction of the tumour, a distinct decrease of urinary high molecular weight forms was observed. Thus, some macromolecular growth factors seem to be associated with epithelial ovarian carcinomas.

Estrogen and androgen receptor mediated stimulation and inhibition of proliferation by androst-5-ene-3 beta,17 beta-diol in human mammary cancer cells.

Androst-5-ene-3 beta,17 beta-diol (ADIOL) and 5 alpha-androstane-3 beta,17 beta-diol (5 alpha A), which are metabolites of dehydroepiandrosterone and dihydrotestosterone, are known to have estrogenic properties. This study reevaluates the estrogenic effects of ADIOL and 5 alpha A in MCF-7 cells and demonstrates additionally androgen-like inhibitory properties of these compounds in human hormone-dependent mammary cancer cells. ADIOL and 5 alpha A (10-100 nM) stimulate the proliferation of estrogen-sensitive MCF-7 cells. Binding assays with the estrogen receptor and inhibition of stimulation with the antiestrogen tamoxifen support the involvement of the estrogen receptor. On the other hand, the mammary cancer cell line MFM-223 is strongly inhibited by ADIOL and 5 alpha A in the same concentration range. This cell line is androgen receptor positive and is inhibited by androgens, but unresponsive to estrogens and progestins. The inhibitory effects of ADIOL and 5 alpha A in MFM-223 cells are mediated by the androgen receptor as demonstrated by receptor studies and competition experiments with hormone antagonists. ADIOL and 5 alpha A thus possess estrogen- and androgen-like properties and can stimulate or inhibit proliferation of human mammary cancer cells. The reactions of mammary cancer cells to these steroids depend on the receptor content and the growth properties of the individual cell line.

Medroxyprogesterone acetate inhibits the proliferation of estrogen- and progesterone-receptor negative MFM-223 human mammary cancer cells via the androgen receptor.

This study demonstrates for the first time, that medroxyprogesterone acetate (MPA) inhibits the proliferation of the estrogen and progesterone receptor negative mammary cancer cell line MFM-223 via the androgen receptor. MPA is a progestin, which is used in the hormonal treatment of disseminated breast cancer. It binds to the progesterone, androgen, and glucocorticoid receptor and may exert its antiproliferative effects via different receptors. MFM-223 human mammary cancer cells contain a very high level of androgen receptors (160 fmol/mg protein) and low levels of estrogen, progesterone, and glucocorticoid receptors (< 20 fmol/mg protein). This cell line provides therefore a good model system to analyze the possible role of the androgen receptor in the action of MPA avoiding interference with other steroid hormone receptors. Effective inhibition of proliferation is achieved by 10 nM MPA or 1 nM of the androgen dihydrotestosterone, corresponding well to the binding affinities of both compounds (3.6 and 0.18 nM, respectively). The involvement of the androgen receptor was confirmed by competition experiments with antiandrogens. Furthermore, MFM-DHT cells, which are an androgen resistant subline of MFM-223 cells, are also resistant to MPA. This data supports the involvement of the androgen receptor in the action of MPA and additionally rules out direct hormone-independent cytotoxic effects of MPA.

Regulation of androgen receptor mRNA and protein level by steroid hormones in human mammary cancer cells.

The regulation of the human androgen receptor (AR) by steroid hormones in human mammary cancer cells was investigated using immunocytochemical and ligand binding assays for its protein and Northern blot analyses for the corresponding mRNA. MFM-223 cells contain high levels of ARs and are growth-inhibited by dihydrotestosterone (DHT). The AR protein is down-regulated to 57% of the control by 10 nM DHT after 24 h, and the corresponding mRNA is also reduced. The nonsteroidal antiandrogen hydroxyflutamide had no effect on the AR level, whereas after incubation with 1 microM cyproterone acetate a slight down-regulation was observed. The AR level was restored completely after release from a 7 day treatment with DHT. However, only 60% of the control level was restored, if the cells wer grown in the presence of DHT for 6 weeks. In androgen-pretreated cells the proliferation rate remained decreased even after the withdrawal of DHT. Concomitantly the distinct growth inhibition was lost. Transfection experiments demonstrated a reduced activity of the residual androgen receptor in these pretreated cells. In addition to the AR, EFM-19 cells also contain significant amounts of estrogen and progesterone receptors. EFM-19 cells are not growth inhibited by physiological concentrations of DHT. Autoregulation of AR was also found in this cell line. Additionally, reduced levels of AR protein and mRNA were found in EFM-19 cells after treatment with the synthetic progestin R5020. The maximum effect of R5020 was observed at the high concentration of 1 microM. Estrogen treatment with 10 nM 17 beta-estradiol for 3 days reduced the AR level only by 25%.