Blood-brain barrier in vitro models as tools in drug discovery: assessment of the transport ranking of antihistaminic drugs.

In the course of our validation program testing blood-brain barrier (BBB) in vitro models for their usability as tools in drug discovery it was evaluated whether an established Transwell model based on porcine cell line PBMEC/C1-2 was able to differentiate between the transport properties of first and second generation antihistaminic drugs. First generation antihistamines can permeate the BBB and act in the central nervous system (CNS), whereas entry to the CNS of second generation antihistamines is restricted by efflux pumps such as P-glycoprotein (P-gP) located in brain endothelial cells. P-gP functionality of PBMEC/C1-2 cells grown on Transwell filter inserts was proven by transport studies with P-gP substrate rhodamine 123 and P-gP blocker verapamil. Subsequent drug transport studies with the first generation antihistamines promethazine, diphenhydramine and pheniramine and the second generation antihistamines astemizole, ceterizine, fexofenadine and loratadine were accomplished in single substance as well as in group studies. Results were normalised to diazepam, an internal standard for the transcellular transport route. Moreover, effects after addition of P-gP inhibitor verapamil were investigated. First generation antihistamine pheniramine permeated as fastest followed by diphenhydramine, diazepam, promethazine and second generation antihistaminic drugs ceterizine, fexofenadine, astemizole and loratadine reflecting the BBB in vivo permeability ranking well. Verapamil increased the transport rates of all second generation antihistamines, which suggested involvement of P-gP during their permeation across the BBB model. The ranking after addition of verapamil was significantly changed, only fexofenadine and ceterizine penetrated slower than internal standard diazepam in the presence of verapamil. In summary, permeability data showed that the BBB model based on porcine cell line PBMEC/C1-2 was able to reflect the BBB in vivo situation for the transport of antihistaminc drugs and to distinguish between first and second generation antihistamines.

Serum-derived immunoglobulins alter amyloid beta transport across a blood-brain barrier in vitro model.

Since passive immunization with serum-derived immunoglobulins (intravenous immunoglobulins) showed several positive effects in some patients with Alzheimer's disease (AD), intravenous immunoglobulins (IVIG) are discussed as a possible treatment option. IVIG, an antibody product derived from human plasma, contains natural antibodies against amyloid beta(Abeta) peptide. Until now it is not known, how IVIG interferes with pathogenesis in AD, but several proposed mechanisms are in discussion. Receptor types which are involved in transport processes at the BBB are LRP, RAGE and hFcRn. We were looking for an in vitro BBB model expressing these receptors and studied the alteration of transport of Abeta peptides across this model under the influence of immunoglobulins. Cell line ECV304 was found to be suitable for our experiments. We found evidence for involvement of an improved clearance of Abeta across the BBB as well as a decreased Abeta influx from blood to the brain probably following complex formation of immunoglobulins with free Abeta in the periphery. Furthermore, we were able to confirm the activity of IVIG preparations which acted the same way but showed slightly less efficacy in comparison to monoclonal anti-Abeta antibodies. Based on these results we suggest multiple mechanisms responsible for the efficacy of immunotherapy in Alzheimer's disease.

Characterization of two blood-brain barrier mimicking cell lines: distribution of lectin-binding sites and perspectives for drug delivery.

In the present study plant lectins with distinct sugar specificities were applied to two blood-brain barrier (BBB) mimicking cell lines, namely human ECV304 and porcine brain microvascular endothelial cells PBMEC/C1-2 in order to elucidate their glycosylation pattern and to evaluate the lectin-cell interaction for lectin-mediated targeting. The bioadhesive properties of fluorescein-labeled lectins were investigated with monolayers as well as single cells using fluorimetry and flow cytometry, followed by confirmation of the specificity of binding. For PBMEC/C1-2 layers highest binding capacity was found for wheat germ agglutinin (WGA), followed by Dolichus biflorus agglutinin (DBA) whereas single cell experiments revealed a predominance of DBA only. Analyzing ECV304 monolayers and single cells, WGA yielded the strongest interaction without any changes during cultivation. The binding capacities of the other lectins increased significantly during differentiation. As similar results to primary cells and brain sections were observed, both cell lines seem to be suitable as models for lectin-interaction studies. Thus, an additional focus was set on the mechanisms involved in uptake and intracellular fate of selected lectins. Cytoinvasion studies were performed with WGA for human ECV304 cells and WGA as well as DBA for PBMEC/C1-2 cells. For both lectins, the association rate to the cells was dependent on temperature which indicated cellular uptake.

Alteration of the glycocalyx of two blood-brain barrier mimicking cell lines is inducible by glioma conditioned media.

Recent studies showed that glioma conditioned medium is able to induce blood-brain barrier properties in in vitro models. In this regard, it was investigated whether glioma conditioned medium can also influence the lectin-binding capacity of blood-brain barrier in vitro models. For the presented study cell lines PBMEC/C1-2 and ECV304 were chosen because it was previously shown that glioma conditioned medium was able to induce specific blood-brain barrier properties in these cell lines. Six different plant lectins (WGA, STL, LCA, UEA-I, DBA, PNA) with distinct sugar specificities were applied in order to elucidate the glycosylation patterns of cell line PBMEC/C1-2 and ECV304. Lectin-binding studies were carried out with monolayers as well as with single cells. In the case of PBMEC/C1-2 monolayers, results showed a significant increase of the binding of lectins WGA, STL, UEA-I, DBA and PNA after application of 25 pmol lectin when cultured in media containing soluble factors derived from glioma cell line C6, whereas the binding capacity for LCA remained similar. For ECV304 monolayers, a significant decrease of WGA, STL and LCA was observable, whereas UEA-I binding increased in comparison to cells grown in the corresponding basal growth medium without soluble C6 factors. Single cell studies showed less significant, but similar changes in the lectin-interactions with the cell surfaces. In conclusion, it was shown that soluble factors derived from glioma cell line C6 can modulate the "glycocalyx" of blood-brain barrier mimicking cell lines.

Applying blood-brain barrier in vitro models to study the influence of drugs on endothelial cells--effects of selected COX-inhibitors.

The influence of three cyclooxygenase (COX) inhibitors (indometacin, lornoxicam and celecoxib) with different COX-1/COX-2 profiles on endothelial cells using in vitro blood-brain barrier (BBB) models was investigated. For the experiments two BBB mimicking cell lines (PBMEC/C1-2 and ECV304) and primary human umbilical vein endothelial cells (HUVEC) were used. In preliminary tests the two cell lines and HUVEC were characterized by cell ELISA in respect of the presence of the tight junction proteins occludin and zonula occludens protein-1 (ZO-1), the adhesion molecules ICAM-1 and VCAM-1 and the endothelial marker von Willebrand factor (vWF). Then, the influence of indometacin, lornoxicam and celecoxib on the expression of occludin, ZO-1, ICAM-1 and vWF of the two cell lines and HUVEC was analysed by cell ELISA. The COX inhibitors caused an effect on PBMEC/C1-2 and HUVEC but no influence was observed on ECV304. The results of PBMEC/C1-2 and HUVEC indicated that in comparable therapeutical concentrations celecoxib had a higher potential to impair endothelial cells and to decrease the expression of occludin, ZO-1 and ICAM-1 than indometacin and lornoxicam.

A psychosocial analysis of women planning birth outside hospital.

Most of the women requesting out-of-hospital delivery considered delivery a natural process, not an illness requiring hospital care. The women cited freedom of choice concerning the delivery, less anxiety in the home than in the hospital environment, a more personal relationship with the midwife, and, as far as possible, making do without medical equipment. The interviewed women were a selected collective regarding age, parity, socioeconomic status and obstetric risk profile. Nonetheless, the results suggest ways that in-hospital obstetrics can be adapted to meet the requirements of pregnant women. Individualized, family-oriented obstetrics with judicious use of medical technology should be possible in the clinical setting.

p21(waf) correlates with DNA replication but not with prognosis in invasive breast cancer.

BACKGROUND/AIMS: p21(waf) plays a central role both in the regulation of the cell cycle and in DNA replication. Accordingly, p21(waf) is a putative tumour suppressor. The role of p21(waf) expression in breast cancer is still unclear, particularly with respect to the clinical situation. Therefore, this retrospective study was designed to investigate the value of immunohistochemically detected p21(waf) expression in invasive breast cancer. METHODS: Cellular expression of p21(waf) was assessed in 307 breast cancer tissues by immunohistochemistry using the monoclonal antibody, clone 4D10. The data were correlated to established and functional factors of prognosis (age, menopausal status, tumour size, nodal status, tumour grade, receptor status, proliferating cell nuclear antigen (PCNA) expression, Her-2/neu expression, and p53 expression), and to clinical follow up (median observation time, 82 months). RESULTS: Ninety nine of 307 (32.2%) tumour tissues were considered p21(waf) positive (nuclear staining). In the entire study group, p21(waf) expression correlated only with increased PCNA expression (chi(2) test: p = 0.029), and with none of the other investigated markers. In node negative patients (n = 134), p21(waf) expression correlated with increased tumour size and increased PCNA expression, whereas the node positive subgroup (n = 161) showed no correlation with these parameters (lymphonodectomy was done in 295 women). With respect to clinical outcome, p21(waf) expression showed a definite favourable trend in both subgroups (N0: p21(waf) negative, 23 of 87; p21(waf) positive, nine of 43. N+: p21(waf) negative, 63 of 107; p21(waf) positive, 23 of 52), but this observation was not significant (p > 0.05). Multivariate analysis for disease free survival as indicated by Cox regression analysis included all factors investigated. The most striking parameters were nodal status (relative risk (RR), 1.74; p = 0.00001), receptor status (RR, 0.59; p = 0.0085), tumour size (RR, 1.42; p = 0.02), and Her2/neu expression (RR, 1.56; p = 0.033). p21(waf) expression was not significant in the multivariate analysis (p > 0.05). CONCLUSIONS: p21(waf) expression is an independent factor but fails to be of prognostic or predictive value in multivariate analysis. These data confirm the hypothesis of a p53 independent p21(waf) induction and suggest a functional role in the inhibition of PCNA mediated DNA replication.

[Risk of uterine rupture after cesarean section--analysis of 1,086 births]. / Das Risiko der Uterusruptur nach vorausgegangenem Kaiserschnitt--eine Analyse von 1,086 Geburten.

OBJECTIVE: In the presented paper, obstetrical management after previous caesarian section was studied in a large patient collective at the University Department of Gynaecology and Obstetrics in Cologne from 1979 to 1995. Particular attention was given to the feared complication of rupture of the uterus. PATIENTS: From a total of 15,166 deliveries, 1,086 of the births had been preceded by one or more caesarian section. These 1,086 births formed the patient collective for the present study. RESULTS: Vaginal delivery was attempted in 44.5% of patients and was successful in 86% of those cases. If there had been a previous caesarian section, the percentage shifted in favour of vaginal delivery. All patients with more than two previous caesarian sections were delivered by a primary caesarian section. The feared complication of rupture of the uterus occurred in four cases, for which case reports are presented. In view of such cases, signs of imminent uterus rupture often constitute an indication for primary (11.5%) or secondary resectioning (31.9%). No relationship was found between fetal outcome and mode of delivery. CONCLUSION: This retrospective study confirms the general recommendation of vaginal delivery following previous caesarian section as long as risks are minimized by a readiness to proceed with resectioning when signs of imminent rupture of the uterus arise.

The risk of rupture of the uterus: an analysis of 1086 births after previous caesarean section.

In the work presented here, obstetric management after a previous caesarean section was studied in a large patient group at the University Department of Gynecology and Obstetrics in Cologne from 1979 to 1995. Particular attention was given to the feared complication of uterine rupture. From a total of 15 166 deliveries, 1086 of the births had been preceded by one or more caesarean sections. These 1086 births formed the basis for the present study. Vaginal delivery was attempted in 44.5% of patients and was successful in 86% of those cases. Where there had been only one previous caesarean section, the percentage shifted in favor of vaginal delivery. All patients with more than two previous caesarean sections were delivered by elective caesarean section. The feared complication of rupture of the uterus occurred in four cases, for which case reports are presented. In view of such cases, signs of imminent uterus rupture often constitute an indication for elective (11.5%) or emergency resectioning (31.9%). No relationship was found between fetal outcome and mode of delivery. This retrospective study confirms the general recommendation and safety of vaginal delivery after a previous caesarean section as long as risks are minimised by a readiness to proceed with a repeat caesarean when signs of imminent rupture of the uterus arise.

[Bilateral ovarian metastases of intestinal adenocarcinoma as an accidental result in macroscopic inconspicuous ovaries - a case report]. / Bilaterale Ovarialmetastasierung eines gastralen Adenokarzinoms als Zufallsdiagnose bei makroskopisch unauffälligen Ovarien.

We here report the incidental histological diagnosis of bilateral ovarial metastasis of a moderately differentiated adenocarcinoma. The patient involved had undergone a vaginal hysterectomy with frontal and rear colporrhaphy on account of uterine and vaginal prolapse, and had requested a further "prophylactic adnexectomy". During the subsequent search for the primary tumor, a localised gastric carcinoma was diagnosed, concealed in the antrum.

[Treatment of perioperative anxiety in suspected breast carcinoma with a phytogenic tranquilizer]. / Zur Behandlung perioperativer Angste bei Mammakarzinomverdacht mit einem Phytotranquilizer.

OBJECTIVE: The present study examined the anxiolytic effect of the herbal preparation Kavosporal forte in 20 patients with situationally induced anxiety. MATERIAL AND METHODS: The degree of anxiety was acute in that the patients were waiting for the results of a histopathological diagnosis, carried out on account of suspect mammary findings, and therefore feared they were suffering from a mammary carcinoma. RESULTS: A significant reduction of anxiety compared with the placebo control was seen after a week's treatment with Kavosporal forte, levels of anxiety being measured a priori from the combined scores of two self-rating scales and one observer-rated scale. In addition, a significant increase was noted in alertness and a lessening of fatigue, introverted behavior and excitability as well as a reduction in levels of depression under the real therapeutic agent over the observation period. In none of the cases examined did any undesirable side effects occur, and the overall tolerance was also consistently good. CONCLUSIONS: It could therefore be concluded that the preparation under investigation is well suited of amelioration of the anxiety that arises regularly in connection with a mammary biopsy.

[Horner's syndrome following lumbar epidural anaesthesia--subdural block?]. / Horner-Syndrom bei lumbaler Epiduralanästhesie--Subduraler Block?

A 28 year old primipara (37th gestational week) was scheduled to undergo delivery by caesarean section under epidural anaesthesia. An epidural catheter was easily inserted in the L3/L4 interspace. After a negative aspiration test 5 ml of bupivacaine 0.25% plus adrenaline 1:200,000 were injected and five minutes later 2 x 5 ml of bupivacaine 0.5% plus fentanyl 0.005 mg/ml were given. Ten minutes after the test dose the patient reported warmth and paraesthesia in the right leg and pelvis and numbness in the right periorbital region. The catheter was drawn back 1.5 centimeters and the operation could be performed under regional anaesthesia with supplemental doses of bupivacaine. Sensory level at the end of the operation was Th 4 which decreased continuously in the following two hours. Ocular symptoms (miosis, ptosis, right-sided numbness) as well as numbness in the right hand and leg persisted four hours longer. The most probable explanation for this peripheral Horner's syndrome is a subdural blockade caused by the first bupivacaine doses. The only known prophylaxis is a fractioned epidural injection of local anaesthetics.

[Pregnancy-saving measures in premature rupture of fetal membranes in the 22nd week of pregnancy]. / Schwangerschaftserhaltende Massnahmen bei vorzeitigem Blasensprung in der 22. SSW.

The obstetric-perinatologic problems raised by premature rupturing of the membranes in the 22nd gestational week is presented in the form of a case report from the Perinatal Centre of the University of Cologne. By close ultrasonic monitoring of the course of development, measurement of parameters of inflammation and administration of prophylactic antibiotics pregnancy could be prolonged by 9 weeks with good fetal outcome.

[Blind prophylactic antibiotic administration in premature rupture of fetal membranes]. / Die ungezielte prophylaktische Antiobiotikagabe bei vorzeitigem Blasensprung.

In a retrospective study based on patients treated at the Department of Gynecology and Obstetrics of the University of Cologne during 1984-1993, we tested whether antepartal prophylactic application of antibiotics in patients with premature rupture of membranes (PROM) decreases the frequency of maternal or fetal infections. With raising intervals between PROM and birth exceeding 24 hours, the frequency of maternal/fetal infections raised in the total study population (n = 940). The maternal infection rate increased from 4% to 11% and the frequency of fetal sepsis from 4% to 19% with a PROM to birth interval exceeding 24 hours. In the group of patients with prophylactic application of antibiotics the frequency of maternal or fetal infections was not lower than in the group without antibiotics. This holds true for the total study population as well as for subgroups with different gestational ages. In the total study population 12% of antibiotics-treated patients and 22% of their newborns had peripartal infections, whereas in the untreated population 3% of the mothers and 4% of the newborns showed signs of infection. The prophylactic application of antibiotics raises significantly the latency period in combination with or without tocolysis in PROM during 25. and 37. week of gestation.

[Trauma in the last trimester of pregnancy]. / Trauma im letzten Trimenon der Gravidität.

Although accidents during pregnancy are fairly rare, besides endangering the mother, they nearly always mean a vital threat to the fetus: lethality rates are up to 8% for the mother and up to 34% for the fetus. Besides depression of the circulatory system in the mother, coupled with fetal hypoxy, injury to the placenta and uterus is also possible. Moreover, the unborn child may be injured by direct trauma. If the fetus sustains a direct injury, the head of the child is affected in the majority of instances. In addition, the risk of an intrauterine death is very high. If the injured baby survives after a section, permanent damage must be taken into account. Pregnant women who are injured in an accident should quickly be checked by sonography and cardiotocography. If no danger is expected for the child, the usual therapeutic rules that apply for traumatology should be followed. If the mother and child are endangered, a cesarean section must be undertaken along with simultaneous accident-related surgery on the mother. Case reports are presented on three cases in our clinic last year, and the current literature is discussed. All three mothers and newborns survived because of the cooperation between surgeons, gynecologists and pediatricians.

[Isolated ovarian metastasis of primary gallbladder carcinoma with estrogen and androgen production]. / Isolierte Ovarialmetastase eines primären Gallenblasenkarzinoms mit Ostrogen- und Androgenproduktion.

The rare case of an isolated ovarian metastasis of a primary adenocarcinoma of the gallbladder is reported: the ovarian metastasis was associated with androgen and estrogen production. The differential diagnoses are discussed.

Immunohistochemical detection of epidermal growth factor receptor lacks prognostic significance for breast carcinoma.

OBJECTIVE: We sought to determine whether the immunohistochemical detection of epidermal growth factor receptor (EGF-R) in primary cancer tissues is of prognostic significance in patients with breast carcinoma. METHODS: Paraffin-embedded tissues from 244 study subjects with primary breast carcinomas were tested immunohistochemically for the presence of EGF-R and were compared in a retrospective study with clinical outcome. RESULTS: Epidermal growth factor receptor was detected in the tumors of 49 (20.1%) of the 244 study subjects. The incidence of EGF-R detection was comparable in subjects with disease-free lymph nodes (T1-4, N0, M0, n = 111; EGF-R present 22.5%) or those whose nodes contained carcinoma (T1-4, N+, M0, n = 133; EGF-R present 18.9%). No reliable correlation was found in either group between EGF-R detection and clinical, functional, or morphologic prognostic indicators that included age, menopausal status, tumor size, tumor grade, nodal status, and hormone receptor status. Relapse-free survival and overall survival (median observation time 62.5 months) did not differ between patients with EGF-R-positive or EGF-R-negative breast carcinoma specimens. CONCLUSIONS: In our experience, the immunohistochemical determination of EGF-R in routine formalin-fixed, paraffin-embedded tumor specimens fails to provide useful information concerning the prognosis of patients with primary breast carcinoma.

[An consultation concept for integrating psychosomatic medicine in gynecology]. / Ein praxisnahes Konzept zur Integration psychosomatischer Medizin in der Gynäkologie.

This paper is meant to survey the results of an interdisciplinary concept of psychosomatic patient treatment carried out for two years at the Department of Gynaecology and Obstetrics of the University of Cologne. This integral programme offers a psychosomatic consultation hour taking place once a week at the Department of Gynaecology and Obstetrics. Each first consultation is attended both by a psychosomatic specialist and a gynaecologist. The hypothesis underlying this new programme, which supposes that the integration of the psychosomatic treatment into the everyday routine of the clinic would make it easier for the patients to find access to psychological help, has proved right in view of the experience made so far. In comparison to the former counselling concept, the newly established gynaecologic-psychosomatical programme has increased the attendance by 300%. Thanks to the integrated consultation hour the way of seeing gynaecological problems in a psychosomatic context has increased considerably within the clinic itself so that these problems can now often be solved directly between gynaecologist and patient without making use of the offered consultation hour. Experiences made so far prove that the psychosomatic consultation hours are a valuable contribution to the diagnostic and therapeutic spectrum of the Department of Gynaecology and Obstetrics.

Prognostic factors for preoperative consultation of women desiring sterilization: findings of a retrospective analysis.

In this exploratory study, 37 sterilized women applying for sterilization reversal were questioned thoroughly to establish why they had decided to undergo sterilization and why they now wished for it to be reversed. Taking an interactive behavioral model as our starting point, we concentrated on the psychosocial circumstances leading to the definitive decision to be sterilized. A relationship crisis at the time of sterilization was found to be a prognostically unfavorable factor. Furthermore, 20 of the 37 patients cited new partnerships as their main reason for seeking reversal. Those who felt pressurized by their gynecologist or partner into undergoing sterilization had significantly more problems in overcoming the psychological stress accompanying such an operation than those who, through a series of consultations on contraception, had had sufficient time and opportunity to make their own decisions. Sterilization performed for medical reasons was found to have particularly problematical consequences, especially where the doctor had made the decision largely on his own, failing to give an adequate explanation for the medical necessity of the operation. Regarding the time chosen for sterilization, the study revealed that the patient's postoperative psychological condition was significantly worse when sterilization was carried out immediately after a delivery, after abortion or after Caesarean section, rather than in the interval between pregnancies. The resulting increase in the incidence of psychosomatic complaints and depressive states is also confirmed in the literature. The findings of this study offer practical suggestions for improved preoperative consultation and should help to determine the course of action to be taken when a patient wishes to be sterilized.

PIP: 37 sterilized women who requested sterilization reversal at the department of gynecology and obstetrics at the University of Cologne in Germany completed a questionnaire on why they chose sterilization and later wanted it reversed. The most common reasons for undergoing sterilization were: a crisis in the relationship with their partner (12), social reasons (7), and birth limiting (7). 3 women had been sterilized without their consent. 11 women, including the 3 who were sterilized without their consent, noted that a 3rd person (partner, physician, mother, girlfriend) was the decisive factor leading to sterilization. Key reasons for applying for sterilization reversal included a new relationship (20), renewed desire to have children with original partner (9), and death of a child (5). 23 of the women had undergone postpartum sterilization. They had a greater feeling of having been manipulated by a 3rd person than those who underwent interval sterilization. Only 11 of the 37 women had wanted their last child before sterilization. 65% of all the women sought counseling from a gynecologist about sterilization. The gynecologist apparently influenced the final decision in about 33% of women. In 60% of cases, the women had gone through with the sterilization without the full consent of their partners. 15 women had suffered from depression requiring drug therapy. 18 women believed that the sterilization produced a negative psychosocial situation, especially among postpartum sterilization cases and those who felt they were coerced to undergo sterilization. Sterilization appeared to have a positive effect on sexual well-being. These findings confirm the importance of extensive preoperative consultation, ideally with both partners, and of the need for the woman to make her own decision without any manipulative influence from a third party.

[Experiences with definitive contraception--results of a follow-up study of sterilized women]. / Erfahrungen mit der definitiven Kontrazeption--Ergebnisse einer Follow-up-Untersuchung sterilisierter Frauen.

Within the framework of a retrospective analysis, 96 patients who had undergone sterilisation at the Department of Gynaecology at the University of Cologne between 1988 and 1991, had been questioned as to whether they were content about their former decision in favour of a definitive contraception. At present, 94% of these women fully agreed with their former decision. The circumstances influencing the decision for sterilisation before the operation i.e. age, number of children, motivation, situation between partners, date of operation and medical consultation - were compared to those of a number of sterilised women seeking refertilization. We want to stress the importance of a sufficiently long decision process, accompanied by individual medical consultation.