Olaf Buchweitz

Increased expression of the adult stem cell marker Musashi‐1 in endometriosis and endometrial carcinoma

Adult stem cells are thought to be responsible for the high regenerative capacity of the human endometrium, and have been implicated in the pathology of endometriosis and endometrial carcinoma. The RNA‐binding protein Musashi‐1 is associated with maintenance and asymmetric cell division of neural and epithelial progenitor cells. We investigated expression and localization of Musashi‐1 in endometrial, endometriotic and endometrial carcinoma tissue specimens of 46 patients. qPCR revealed significantly increased Musashi‐1 mRNA expression in the endometrium compared to the myometrium. Musashi‐1 protein expression presented as nuclear or cytoplasmic immunohistochemical staining of single cells in endometrial glands, and of single cells and cell groups in the endometrial stroma. Immunofluorescence microscopy revealed colocalization of Musashi‐1 with its molecular target Notch‐1 and telomerase. In proliferative endometrium, the proportion of Musashi‐1‐positive cells in the basalis layer was significantly increased 1.5‐fold in the stroma, and three‐fold in endometrial glands compared to the functionalis. The number of Musashi‐1 expressing cell groups was significantly increased (four‐fold) in proliferative compared to secretory endometrium. Musashi‐1 expressing stromal cell and cell group numbers were significantly increased (five‐fold) in both endometriotic and endometrial carcinoma tissue compared to secretory endometrium. A weak to moderate, diffuse cytoplasmic glandular staining was observed in 50% of the endometriosis cases and in 75% of the endometrioid carcinomas compared to complete absence in normal endometrial samples. Our results emphasize the role of Musashi‐1‐expressing endometrial progenitor cells in proliferating endometrium, endometriosis and endometrioid uterine carcinoma, and support the concept of a stem cell origin of endometriosis and endometrial carcinoma. Copyright

The adult stem cell marker Musashi-1 modulates endometrial carcinoma cell cycle progression and apoptosis via Notch-1 and p21WAF1/CIP1

The RNA‐binding protein Musashi‐1 has been proposed to maintain stem cell function during development and regenerative processes as a modulator of the Notch‐1 signaling pathway. Musashi‐1 expression is upregulated in endometrial carcinoma, however, its pathogenetic role in this tumor entity is unknown. Here we investigate the functional impact and mode of action of Musashi‐1 on endometrial carcinoma cell behaviour in vitro. Aldehyde dehydrogenase‐1 activity and side population (SP) measurement by Hoechst dye exclusion revealed that the Ishikawa endometrial carcinoma cell line contains a pool of putative cancer stem cells. Musashi‐1 expression is 20.8‐fold upregulated in SP+ compared to SP‐ and equally distributed between ALDH+ and ALDH‐ cell pools. siRNA‐mediated knockdown of Musashi‐1 mRNA expression lead to an altered expression of the signaling receptor Notch‐1 and its downstream targets, the transcription factor Hes‐1 and the cell cycle regulators p21WAF1/CIP1 and cyclin B1, as determined by Western blotting and quantitative real‐time PCR. Flow cytometric and ELISA analyses revealed that Musashi‐1‐mediated modulation of these factors exerted an antiproliferative effect on the cell cycle, and increased apoptosis in endometrial carcinoma cells. We conclude that Ishikawa cells contain a subpopulation of cells with stem cell‐like properties. Musashi‐1 modulates endometrial carcinoma cell cycle progression and apoptosis via the stemness‐related factors Notch‐1, Hes‐1 and p21WAF1/CIP1, thus emerging as a novel future target for endometrial carcinoma therapy.

Prospective evaluation of the systemic immune response following abdominal, vaginal, and laparoscopically assisted vaginal hysterectomy

BackgroundAlterations in serum levels of cytokine interleukin-6 (IL-6) and acute-phase protein C-reactive protein (CRP) correlate directly with extent of tissue damage and inflammatory reaction. We therefore prospectively compared the postoperative levels of IL-6 and CRP following abdominal (AH), vaginal (VH), and laparoscopically assisted vaginal hysterectomy (LAVH).MethodsA total of 29 patients were included in the study (10 VH, 10 LAVH, 9 AH). Nine blood samples were taken from each patient at various time points before, during, and after surgery. CRP and IL-6 were measured under standardized conditions using ELISA and turbidometry.ResultsPreoperative levels of IL-6 and CRP were low in all three patient groups. There was a significant increase in the IL-6 level in patients undergoing AH at the time of peritoneal closure that reached a maximum 2 h postoperatively and remained significantly elevated for 12 h postoperatively when compared to the IL-6 levels of patients undergoing VH or LAVH (p<0.05). The levels of the IL-6 time courses differed significantly among the three operative procedures (p=0.013). In contrast, the levels of the CRP time courses did not differ significantly (p=0.066); however, CRP expression was elevated 36 h postoperatively in patients undergoing AH, as compared with those undergoing VH.ConclusionElevated IL-6 levels subsequent to AH may reflect significantly greater tissue damage in these patients than in patients who undergo VH or LAVH. LAVH should therefore be considered in cases that cannot be managed by the vaginal route alone.

Chromosomal losses of regions on 5q and lack of high-level amplifications at 8q24 are associated with favorable prognosis for ovarian serous carcinoma.

In this study, we describe characteristic chromosomal alterations in a consecutive series of 96 serous ovarian tumors by comparative genomic hybridization. We analyze their association with different pathways of progression, histological grade, and clinical outcome. The most striking difference between low‐grade and high‐grade serous carcinomas was seen in a higher incidence of chromosomal gains at 3q and 20q and losses of 13q in the high‐grade carcinomas. In addition, high‐level amplifications were significantly more frequent in high‐grade carcinomas, specifically involving regions on 3q and 8q. Chromosomal amplifications of 19p and 19q and losses of 4q and 5q were among the most frequent changes found in both low‐grade and high‐grade carcinomas, distinguishing them from borderline tumors, which had very few recurrent alterations. The most significant impact on survival of patients with invasive carcinomas Stage II–IV was observed for high‐level amplifications of regions on 8q (mean overall survival (OS) 69 versus 27 months, P = 0.0006). Interestingly, low‐level gains on 8q do not show any impact compared to cases with no alteration. Surprisingly, chromosomal losses on 5q had a protective impact (mean OS 36 versus 76 months, P = 0.0007). Combination of both parameters resulted in two risk groups. Low risk: loss on 5q, no amplification on 8q (mean OS 84 months); high risk: no loss on 5q, amplification on 8q (mean OS 26 months). This difference is even more pronounced, if only cases with residual tumor of less than 2 cm are included, resulting in a 5‐year survival of 100% and 0% (P = 0.0005).

Laparoscopic management of tubo-ovarian abscesses: retrospective analysis of 60 cases.

BackgroundThe laparoscopic management of tubo-ovarian abscesses (TOA) was evaluated. The study sought to answer the following question: Does operative laparoscopy with only incision of the abscess cavity and lavage (organpreserving treatment) improve intraoperative and postoperative safety and long-term prospects of fertility as compared with laparoscopic salpingectomy or salpingo-oophorectomy (ablative treatment)?MethodsA retrospective chart review of 60 patients with TOA undergoing laparoscopic treatment in combination with broad-spectrum antibiotics from 1994 to 1998 was performed. Patients not wishing to have children underwent salpingectomy or salpingo-oophorectomy, whereas patients wishing to remain fertile were treated by means of an organpreserving procedure. To investigate the operative and reproductive outcome, patients were interviewed by telephone.ResultsOf 60 women with TOA, 25 were treated laparoscopically, preserving the internal genital organs, and 35 underwent ablative treatment. Apart from one postoperative readmission because of lower pelvic pain in the organpreserving group, there were no operative complications or serious systemic sequelae. In contrast, there was a significantly higher incidence of intraoperative and postoperative complications when ablative treatment was performed: one intestinal perforation requiring subsequent laparotomy, four serosal lesions, two lesions of the greater omentum, two lacerated collaterals of the internal iliac artery, one postoperative fever higher than 38°C for 2 days, two bowel obstructions, one thrombosis of the upper leg, and one thrombosis of the lower leg. There were no significant differences between the two patient groups in body mass index, duration of pelvic pain, laboratory findings at admission, ultrasonic assessment of abscess size, and the extent of the abscess at laparoscopy.ConclusionsWhen laparoscopic treatment of TOA is performed, organ-preserving treatment should be chosen irrespective of the patient’s age or desire to have children because of the risk of complications.

Aberrant expression of the pluripotency marker SOX-2 in endometriosis

Expression of the pluripotency factors SOX-2, OCT-4, KLF-4, and NANOG was analyzed by quantitative real-time polymerase chain reaction, immunohistochemistry, and immunofluorescence microscopy in the endometrium, myometrium, and endometriotic tissue of 36 patients. Aberrant expression of SOX-2 may indicate a stem cell origin of endometriosis, whereas the presence of all progenitor markers in endometrial tissue marks the endometrium as a potential source for induced pluripotent stem cell generation.

The Diagnostic Dilemma of Minimal and Mild Endometriosis under Routine Conditions

STUDY OBJECTIVE To evaluate the reliability of diagnosing minimal and mild endometriosis under routine conditions, and to determine to what extent disease activity is taken into account. DESIGN Retrospective analysis (Canadian Task Force classification II-2). SETTING University teaching hospital. INTERVENTION Laparoscopy. PATIENTS One hundred eighteen consecutive women with minimal and mild endometriosis undergoing routine surgery between 1994 and 1999. MEASUREMENTS AND MAIN RESULTS Analytic parameters were the total number of endometriotic lesions; intraoperative description of pigmented, nonpigmented, and nondefined lesions; and number of extirpated lesions and histologic detection rate. In 118 patients, 311 suspected endometriotic lesions were documented. Nonpigmented lesions were reported in only 27% of women. In 51% of surgical reports no importance was attached to disease morphology or activity. Only 1.2 biopsies/patient were taken. The histologic detection rate was 56%. In 49 patients the assumed intraoperative diagnosis was confirmed by histologic examination. CONCLUSIONS Intraoperative description of endometriotic lesions is inadequate. Little attention is paid to the activity of the illness. There is room for improvement in the number of excisions and histologic detection, and an attempt should be made to find a way out of this diagnostic dilemma.

Detection of nonpigmented endometriotic lesions with 5-aminolevulinic acid-induced fluorescence.

STUDY OBJECTIVE To evaluate the feasibility of fluorescence diagnosis of nonpigmented (red and white) endometriotic lesions with 20 mg/kg of 5-aminolevulinc acid (5-ALA) 5-7 and 10-14 hours before surgery. DESIGN Prospective analysis (Canadian Task Force classification II-2). SETTING University hospital. PATIENTS Twenty-four consecutive patients with suspected endometriosis undergoing laparoscopy. INTERVENTION Laparoscopic surgery under white light illumination and fluorescence diagnosis. MEASUREMENTS AND MAIN RESULTS The total number of endometriotic lesions detected under white light illumination was compared with the number of lesions detected with fluorescence diagnosis. Fluorescence diagnosis yielded an overall improvement of 35% in the detection of nonpigmented endometriotic lesions compared with white light illumination. Sensitivity (91%) and specificity (79%) were similar 5-7 and 10-14 hours before surgery. CONCLUSION The dosage of 20 mg/kg body weight of 5-ALA is feasible for fluorescence diagnosis of nonpigmented endometriosis. Sensitivity of fluorescence diagnosis with 20 mg/kg is similar to that achieved with a 30-mg/kg dose. Sensitivity does not change within the application period 5-7 and 10-14 hours before surgery.

A prospective randomized trial of closing laparoscopic trocar wounds by transcutaneous versus subcuticular suture or adhesive papertape

BackgroundSeveral methods for closure of trocar wounds are known in laparoscopic surgery. The choice of technique (mostly transcutaneous or subcuticular suture or adhesive papertape) is often based on the surgeon’s personal experience. Thus, the objective of this trial was to assess the impact of these closure methods on potential complications of wound healing, cosmetic outcome, and patient satisfaction.MethodsSixty patients undergoing operative laparoscopic surgery for gynecologic indications were enrolled in this prospective randomized trial. Five-millimeter port-site incisions were closed either with subcuticular or transcutaneous absorbable sutures (4–0 polyglactin 910) or with adhesive papertape. Postoperative complications, pain, and patient satisfaction with scars were evaluated at 3-month follow-up after operation using a questionnaire.ResultsData from 52 patients who returned the questionnaire were analyzed. Dissatisfying cosmetic results were reported significantly more frequently after subcuticular sutures (p < 0.05). Assessment of patient satisfaction with cosmetic outcome on a visual–analogue scale revealed significantly better results after transcutaneous skin closure than with other approaches (p < 0.05). Adverse wound healing (e.g., infections and dehiscence) was observed most frequently in the subcuticular suture group. Also, the rate of painful scars was highest with this technique.ConclusionsTranscutaneous closure with absorbable suture material seems to be the most suitable technique for closure of laparoscopic port-site incisions.

Endometriose und Aktivität

ZusammenfassungHinter der Vielzahl der Erscheinungsbilder der Endometriose verbergen sich unterschiedlich “aktive” Areale der Erkrankung. Die nicht pigmentierten Läsionen der Endometriose gehen mit einer erhöhten Expression von unterschiedlichen Wachstumsfaktoren und Zytokinen einher. Dadurch unterscheiden sie sich von pigmentierten Läsionen der Endometriose. Setzt man die erhöhte Expression bestimmter Parameter mit “Aktivität” gleich, bedeutet dies, dass die nicht pigmentierten Läsionen aktive Formen der Erkrankung darstellen.Die bisherigen klinischen Resultate berücksichtigen die Aktivität der Erkrankung nicht oder nur marginal. Eine exakte Dokumentation der Aktivität während der diagnostischen/operativen Laparoskopie stellt die Grundvoraussetzung für eine adäquate Therapie dar. Darüber hinaus liefert die Kenntnis über biochemische Eigenschaften der Endometrioseläsionen wichtige Rückschlüsse auf die Pathogenese der Erkrankung und eröffnet so neue Therapieoptionen.AbstractThe numerous morphologic aspects of different endometriotic lesions can be related to different stages of activity of the endometriotic disease. Non-pigmented endometriotic lesions are demonstrating higher levels of growth factors and cytokines than pigmented lesions. It seems that high levels of growth factors (VEGF, angiopoietins, MMP) are related with high biologic activity, therefore the pigmented lesions represent the active manifestation of the disease.Until now most clinical studies have not taken into account the activity of the disease. An exact documentation of the activity during laparoscopy represents a prerequisite for an adequate therapy. Furthermore, the improvement of our knowledge will lead to a better understanding of the pathogenesis of the endometriotic disease and to new options in the treatment of endometriosis.