Visualization of endothelial cell cycle dynamics in mouse using the Flt-1/eGFP-anillin system

Endothelial cell proliferation is a key process during vascular growth but its kinetics could only be assessed in vitro or ex vivo so far. To enable the monitoring and quantification of cell cycle kinetics in vivo, we have generated transgenic mice expressing an eGFP-anillin construct under control of the endothelial-specific Flt-1 promoter. This construct labels the nuclei of endothelial cells in late G1, S and G2 phase and changes its localization during the different stages of M phase, thereby enabling the monitoring of EC proliferation and cytokinesis. In Flt-1/eGFP-anillin mice, we found eGFP⁺ signals specifically in Ki67⁺/PECAM⁺ endothelial cells during vascular development. Quantification using this cell cycle reporter in embryos revealed a decline in endothelial cell proliferation between E9.5 to E12.5. By time-lapse microscopy, we determined the length of different cell cycle phases in embryonic endothelial cells in vivo and found a M phase duration of about 80 min with 2/3 covering karyokinesis and 1/3 cytokinesis. Thus, we have generated a versatile transgenic system for the accurate assessment of endothelial cell cycle dynamics in vitro and in vivo.     

Biliary complications in liver transplantation: Impact of anastomotic technique and ischemic time on short- and long-term outcome

AIM: To elucidate the impact of various donor recipient and transplant factors on the development of biliary complications after liver transplantation.METHODS: We retrospectively reviewed 200 patients of our newly established liver transplantation(LT) program, who received full size liver graft. Biliary reconstruction was performed by side-to-side(SS), end-to-end(EE) anastomosis or hepeaticojejunostomy(HJ). Biliary complications(BC), anastomotic stenosis, bile leak, papillary stenosis, biliary drain complication, ischemic type biliary lesion(ITBL) were evaluated by studying patient records, corresponding radiologic imaging and reports of interventional procedures [e.g., endoscopic retrograde cholangiopancreatography(ERCP)]. Laboratory results included alanine aminotransferase(ALT), gammaglutamyltransferase and direct/indirect bilirubin with focus on the first and fifth postoperative day, six weeks after LT. The routinely employed external bile drain was examined by a routine cholangiography on the fifth postoperative day and six weeks after transplantation as a standard procedure, but also whenever clinically indicated. If necessary, interventional(e.g., ERCP) or surgical therapy was performed. In case of biliary complication, patients were selected, assigned to different complication-groups and subsequently reviewed in detail. To evaluate the patients outcome, we focussed on appearance of postoperative/post-interventional cholangitis, need for rehospitalisation, retransplantation, ITBL or death caused by BC.RESULTS: A total of 200 patients [age: 56(19-72), alcoholic cirrhosis: n = 64(32%), hepatocellular carcinoma: n = 40(20%), acute liver failure: n = 23(11.5%), cryptogenic cirrhosis: n = 22(11%), hepatitis B virus /hepatitis C virus cirrhosis: n = 13(6.5%), primary sclerosing cholangitis: n = 13(6.5%), others: n = 25(12.5%) were included. The median follow-up was 27 mo until June 2015. The overall biliary complication rate was 37.5%(n = 75) with anastomotic strictures(AS): n = 38(19%), bile leak(BL): n = 12(6%), biliary drain complication: n = 12(6%); papillary stenosis(PS): n = 7(3.5%), ITBL: n = 6(3%). Clinically relevant were only 19%(n = 38). We established a comprehensive classification for AS with four grades according to clinical relevance. The reconstruction techniques [SS: n = 164, EE: n = 18, HJ: n = 18] showed no significant impact on the development of BCs in general(all n 0.05), whereas in the HJ group significantly less AS were found(P = 0.031). The length of donor intensive care unit stay over 6 d had a significant influence on BC development(P = 0.007, HR = 2.85; 95%CI: 1.33-6.08) in the binary logistic regression model, whereas other reviewed variables had not [warm ischemic time 45 min(P = 0.543), cold ischemic time 10 h(P = 0.114), ALT init 1500 U/L(P = 0.631), bilirubin init 5 mg/d L(P = 0.595), donor age 65(P = 0.244), donor sex(P = 0.068), rescue organ(P = 0.971)]. 13%(n = 10) of BCs had no therapeutic consequences, 36%(n = 27) resulted in repeated lab control, 40%(n = 30) received ERCP and 11%(n = 8) surgical therapy. Fifteen(7.5%) patients developed cholangitis [AS(n = 6), ITBL(n = 5), PS(n = 3), biliary lesion BL(n = 1)]. One patient developed ITBL twelve months after LT and subsequently needed retransplantation. Rehospitalisation rate was 10.5 %(n = 21) [AS(n = 11), ITBL(n = 5), PS(n = 3), BL(n = 1)] with intervention or reinterventional therapy as main reasons. Retransplantation was performed in 5(2.5%) patients [ITBL(n = 1), acute liver injury(ALI) by organ rejection(n = 3), ALI by occlusion of hepatic artery(n = 1)]. In total 21(10.5%) patients died within the follow-up period. Out of these, one patient with AS developed severe fatal chologenic sepsis after ERCP.CONCLUSION: In our data biliary reconstruction technique and ischemic times seem to have little impact on the development of BCs.     

Ueber congenitale Theilung des Parietale durch eine quere oder schräge Sutur

Ohne ZusammenfassungHierzu Taf. IV. Fig. 1.     

T and B cells target identical regions of the non-collagenous domain 1 of type VII collagen in epidermolysis bullosa acquisita

Epidermolysis bullosa acquisita (EBA) is a severe immunobullous disease and is caused by IgG against type VII collagen (Col VII) of anchoring fibrils. In this study, utilizing ELISA and immunoblot, 13/15 EBA sera but 0/20 bullous pemphigoid sera and 0/30 healthy control sera showed IgG reactivity with distinct recombinant subregions of the non-collagenous domain 1 (NC1) of Col VII. In two EBA patients, IgG titers against Col VII-NC1 were grossly correlated to clinical disease activity. Moreover, Col VII-reactive T cells were identified in a representative EBA patient which recognized identical subdomains of Col VII-NC1. These findings strongly suggest that (1) the Col VII-NC1 ELISA is a powerful tool for making the diagnosis of EBA, (2) Col VII-specific IgG grossly relates to disease activity and (3) IgG reactivity is associated with T cell recognition of identical subdomains of Col VII-NC1.     

Distribution of Langerhans cells and mast cells within the human oral mucosa: new application sites of allergens in sublingual immunotherapy?

Background:  Sublingual immunotherapy (SLIT) represents an alternative to subcutaneous immunotherapy. While antigen-presenting cells such as Langerhans cells (LCs) are thought to contribute to the effectiveness of SLIT, mast cells (MCs) most likely account for adverse reactions such as sublingual edema. As little is known about LCs and MCs within the oral cavity, we investigated their distribution in search for mucosal sites with highest LCs and lowest MCs density.
Methods:  Biopsies were taken simultaneously from human vestibulum, bucca, palatum, lingua, sublingua, gingiva, and skin. Immunohistochemistry and flow cytometry were used to detect MCs, LCs and high affinity receptor for IgE (FcεRI) expression of LCs. Mixed lymphocyte reactions were performed to assess their stimulatory capacity.
Results:  Highest density of MCs was detected within the gingiva, while the lowest density of MCs was found within the palatum and lingua. However, sublingual MCs were located within glands, which might explain swelling of sublingual caruncle in some SLIT patients. Highest density of LCs was detected within the vestibular region with lowest density in sublingual region. Highest expression of FcεRI was detected on LCs within the vestibulum. Furthermore LCs from different regions displayed similar stimulatory capacity towards allogeneic T cells.
Conclusions:  In view of our data, different mucosal regions such as the vestibulum might represent alternative SLIT application sites with potent allergen uptake. Our data might serve as a basis for new application strategies for SLIT to enhance efficiency and reduce local adverse reactions.     

Prevalence of generalized anxiety at eight weeks postpartum

Summary Objective: This study investigated the prevalence and nature of generalized anxiety symptoms in women who were eight weeks postpartum. Method: A community-based sample of 68 postpartum women completed an interview assessing generalized anxiety disorder and depression and a self-report measure of worry associated with concerns relevant to postpartum women. Results: Three women (4.4%) met DSM-IV criteria for generalized anxiety disorder, and an additional 19 women (27.9%) endorsed subsyndromal difficulties with generalized anxiety. Approximately one third of these women endorsed symptoms of depression. In contrast, only two woman met criteria for major depressive disorder. Conclusion: Postpartum generalized anxiety has a higher prevalence than postpartum depression. Received February 26, 2002; accepted November 11, 2002 Published online January 31, 2003 Acknowledgement This work was supported by a New Faculty Scholar Award and a Graduate Research Fellowship from the University of North Dakota. The authors would like to express their appreciation to Jennifer Brendle, Peter Schmutzer, Talia Tweten, and Chad Lystad for their assistance with this research. Correspondence: Amy Wenzel, Ph.D., Department of Psychology, University of North Dakota, Grand Forks, ND 58202-8380, U.S.A.; e-mail: amy_wenzel@und.nodak.edu