bluuebunny/crossref_metadata_2025 huggingface dataset:
Floxed allele for conditional inactivation of the GABAB(1) gene
Authors: Haller Corinne , Casanova Emilio , Müller Matthias , Vacher Claire‐Marie , Vigot Rejan , Doll Thierry , Barbieri Samuel , Gassmann Martin , Bettler Bernhard
Published: October 2004

GABAB receptors are the G‐protein‐coupled receptors for the neurotransmitter GABA. GABAB receptors are broadly expressed in the nervous system. Their complete absence in mice causes premature lethality or—when mice are viable—epilepsy, impaired memory, hyperalgesia, hypothermia, and hyperactivity. A spatially and temporally restricted loss of GABAB function would allow addressing how the absence of GABAB receptors leads to these diverse phenotypes. To permit a conditional gene inactivation, we flanked critical exons of the GABAB(1) gene with lox511 sites. GABAB(1)lox511/lox511 mice exhibit normal levels of GABAB(1) protein, are fertile, and do not display any behavioral phenotype. We crossed GABAB(1)lox511/lox511 with Cre‐deleter mice to produce mice with an unrestricted GABAB receptor elimination. These GABAB(1)−/− mice no longer synthesize GABAB(1) protein and exhibit the expected behavioral abnormalities. The conditional GABAB(1) allele described here is therefore suitable for generating mice with a site‐ and time‐specific loss of GABAB function. genesis 40:125–130, 2004. © 2004 Wiley‐Liss, Inc.
X‐inactivation is stably maintained in mouse embryos deficient for histone methyl transferase G9a
Authors: Ohhata Tatsuya , Tachibana Makoto , Tada Masako , Tada Takashi , Sasaki Hiroyuki , Shinkai Yoichi , Sado Takashi
Published: October 2004

Summary: One of the two X chromosomes becomes inactivated during early development of female mammals. Recent studies demonstrate that the inactive X chromosome is rich in histone H3 methylated at Lys‐9 and Lys‐27, suggesting an important role for these modifications in X‐inactivation. It has been shown that in the mouse Eed is required for maintenance of X‐inactivation in the extraembryonic lineages. Interestingly, Eed associates with Ezh2 to form a complex possessing histone methyltransferase activity predominantly for H3 Lys‐27. We previously showed that G9a is one of the histone methyltransferases specific for H3 Lys‐9 and is essential for embryonic development. Here we examined X‐inactivation in mouse embryos deficient for G9a. Expression of Xist, which is crucial for the initiation of X‐inactivation, was properly regulated and the inactivated X chromosome was stably maintained even in the absence of G9a. These results demonstrate that G9a is not essential for X‐inactivation. genesis 40:151–156, 2004. © 2004 Wiley‐Liss, Inc.
Sox10‐rtTA mouse line for tetracycline‐inducible expression of transgenes in neural crest cells and oligodendrocytes
Authors: Ludwig Andreas , Schlierf Beate , Schardt Anke , Nave Klaus‐Armin , Wegner Michael
Published: October 2004

Using gene targeting, we inserted a high‐affinity variant of the reverse tetracycline controlled transactivator (rtTA) into the genomic Sox10 locus. This rtTA transgene faithfully recapitulated Sox10 expression in the emerging neural crest, several of its derivatives, and in oligodendrocytes. It was furthermore able to induce expression of a tetracycline inducible transgenic reporter gene in a doxycycline‐dependent manner. Induction was fast, with substantial reporter gene expression visible 6 h after the onset of doxycycline treatment. Shut‐off, in contrast, exhibited delayed kinetics, which probably correlated with doxycycline clearance rates. This mouse provides a useful tool for generating tetracycline‐controlled gene expression in neural crest and oligodendrocytes. genesis 40:171–175, 2004. © 2004 Wiley‐Liss, Inc.
NG2 proteoglycan‐expressing cells of the adult rat brain: Possible involvement in the formation of glial scar astrocytes following stab wound
Authors: Alonso G.
Published: October 2004

Stab wound lesion to the adult central nervous system induces strong proliferative response that is followed by the formation of a dense astroglial scar. In order to determine the origin of those astrocytes composing the glial scar, the cell proliferation marker bromodeoxyuridine (BrdU) was administered to lesioned rats that were fixed 3 h or 6 days later. At 3 h after the BrdU administration, labeled nuclei were frequently associated with either NG2+ cells or microglia/macrophages, but rarely with astrocytes expressing glial fibrillary acidic protein (GFAP). Six days later, by contrast, numerous BrdU‐labeled nuclei were associated with astrocytes located along the lesion borders. After the injection of a viral vector of the green fluorescent protein (GFP) into the lesional cavity, GFP was preferentially detected within NG2‐ or GFAP‐labeled cells when lesioned animals were fixed 1 or 6 days after the injections, respectively. The combined detection of glial markers within cells present in the lesioned area indicated that, although they rarely express GFAP, the marker of mature astrocytes, NG2+ cells located along the lesion borders frequently express nestin and vimentin, i.e., two markers of immature astrocytes. Lastly, chronic treatment of lesioned rats with dexamethasone was found to inhibit the proliferation of NG2+ cells present within the lesioned area and to subsequently alter the formation of a dense astroglial scar. Taken together, these data strongly suggest that following a surgical lesion, at least a portion of the astrocytes that constitute the glial scar are issued from resident NG2+ cells. © 2004 Wiley‐Liss, Inc.
Can Turner syndrome teach us about the pathogenesis of chronic cholestasis?
Authors: Milkiewicz Piotr , Heathcote Jenny , Jaeschke Harmut , Mullen Kevin , Moradpour Darius
Published: October 2004

The mechanisms that cause the female predominance of primary biliary cirrhosis (PBC) are uncertain, but the X chromosome includes genes involved in immunological tolerance. We assessed the rate of X monosomy in peripheral white blood cells from 100 women with PBC, 50 with chronic hepatitis C, and 50 healthy controls, by fluorescence in-situ hybridisation. Frequency of X monosomy increased with age in all groups, but was significantly higher in women with PBC than in controls (p<0.0001); age-adjusted back-transformed mean frequencies were 0.050 (95% CI 0.046-0.055) in women with PBC, 0.032 (0.028-0.036) in those with chronic hepatitis C, and 0.028 (0.025-0.032) in controls. We suggest that haploinsufficiency for specific X-linked genes leads to female susceptibility to PBC.
Thrombotic thrombocytopenic purpura: 24 years of experience at the American University of Beirut Medical Center
Authors: Shamseddine Ali , Saliba Teddy , Aoun Elie , Chahal Aref , El‐Saghir Nagi , Salem Ziad , Bazarbachi Ali , Khalil Mazen , Taher Ali
Published: October 2004

Thrombotic thrombocytopenic purpura (TTP) is a hematological syndrome defined by the presence of thrombocytopenia and microangiopathic hemolytic anemia without a clinically apparent etiology. Patients may also suffer from fever in addition to neurological and renal impairment. Treatment should be initiated as soon as possible, otherwise this rare disease can be fatal. The main treatment options include therapeutic plasma exchange, fresh frozen plasma infusion, and adjuvant agents such as steroids and antiplatelet drugs. A search of patient records was carried out at the American University of Beirut Medical Center looking for patients who developed TTP over a 24‐year period extending from 1980 to 2003. Relevant information was collected and analyzed. A total of 47 records were found. All presented with anemia and thrombocytopenia, 83% had neurological symptoms, 61.7% had fever and 34% had renal impairment. All patients were treated with a multimodality regimen including therapeutic plasma exchange, FFP infusion, steroids, antiplatelet agents, vincristine and others. 38 (81%) cases achieved complete remission. Out of these, 12 (31.6%) relapsed and responded to treatment. Patients who did not receive plasma exchange were more likely to relapse (P = 0.032). A second relapse was observed in 6 cases. The overall mortality rate from TTP over 24 years was 21.3%. TTP remains a fatal disease. A high index of suspicion should, therefore, always be present. Treatment options should be further developed and patients should directly be referred to tertiary care centers. J. Clin. Apheresis 19:119–124, 2004. © 2004 Wiley‐Liss, Inc.
Comparative study of plateletpheresis using Baxter CS 3000 plus and Haemonetics MCS 3P
Authors: Patel Ashwin P. , Kaur Amarjit , Patel Vinod , Patel Narendra , Shah Dilip , Kanvinde Sunil , Prajapati Sanjay , Patel Hiral , Rathod Dinesh , Adesara Rashmin , Rani Shubha
Published: October 2004

Platelet concentrates made from cell separators are used more frequently due to less donor exposure and leucodepletion. This retrospective study was done to compare plateletpheresis done on two cell separators: Baxter CS 3000 plus and Haemonetics MCS 3p. Plateletpheresis procedures, done from January 1997 to April 2002, were included in the study. One hundred and seven procedures were done on Haemonetics MCS 3p using SDP protocol, 49 procedures were done on Haemonetics MCS 3p using PLP protocol, and 107 were done on Baxter CS 3000 plus. Pre‐procedure donor's platelet count and haemoglobin were comparable in all the groups. Platelet yield was comparable in PLP (6.44 × 1011 platelets) and SDP (5.27 × 1011) protocols, but significantly less in Baxter (4.05 × 1011 platelets, P < 0.001 for PLP and P < 0.05 for SDP). Efficiency of platelet removal was statistically significantly different in all the groups (P < 0.0001), however it was more in PLP (PLP‐55.02%, SDP‐47.38%, Baxter 38.98%). A significant number of products (19.51%) of Baxter failed to comply platelet count of product ≤2,435 × 109/l compared to 5.13% in PLP and 1.23% in SDP group; 36.96% of units from PLP and 28% from SDP qualified for split products compared to 1.18% of Baxter. PLP protocol of Haemonetics MCS 3p gives better platelet yield compared to Baxter CS 3000 plus and SDP protocol of Haemonetics MCS 3p. J. Clin. Apheresis 19:137–141, 2004. © 2004 Wiley‐Liss, Inc.
Donor tolerance and results of stimulation with G‐CSF alone or in combination with dexamethasone for the collection of granulocytes
Authors: Moog Rainer
Published: October 2004

Stimulation of healthy granulocyte donors allows the collection of therapeutic doses of granulocytes. The stimulation with G‐CSF alone was compared with G‐CSF plus dexamethasone. Blood samples were drawn at baseline, at leukapheresis, and at follow‐up visit. Donors answered a questionnaire to evaluate side effects of the stimulation regimen. The combination of G‐CSF and dexamethasone resulted in higher WBC count than G‐CSF alone (39.4 ± 7.8 vs. 34.8 ± 8.3/nl). Glucose (136 ± 45 mg/dl) and lactate dehydrogenase (195 ± 38) increased significantly after stimulation with G‐CSF plus dexamethasone but returned to baseline levels at the follow‐up visit. Generally, stimulation was well tolerated by the donors. A higher rate of mild bone pain and headache was experienced in donors stimulated with G‐CSF plus dexamethasone than in donors receiving G‐CSF alone. Fatigue and myalgia were reported at similar rates in both groups. A high proportion of the donors stated that they would accept a further stimulation and granulocyte donation. At the follow‐up visit, blood counts and chemistry had returned to normal values. J. Clin. Apheresis 19:115–118, 2004. © 2004 Wiley‐Liss, Inc.
Collection of a mobilized peripheral blood apheresis product from a patient with mucopolysaccharidosis type VII and subsequent CD34+ cell isolation
Authors: Hofling A. Alex , Sands Mark S. , Lublin Douglas M. , Bauer Gerhard , Devine Steven
Published: October 2004

The effectiveness of bone marrow transplantation for lysosomal storage diseases like mucopolysaccharidosis type VII (MPSVII) suggests that a gene therapy strategy targeting autologous hematopoietic progenitor cells could be successful. Given the severe systemic manifestations of MPSVII including storage disease in the bone and bone marrow, it was unclear whether sufficient numbers of hematopoietic progenitor cells (CD34+) could be mobilized into the peripheral circulation and subsequently purified from these patients. As reported here, G‐CSF mobilization and apheresis were successful, providing a product of 4 × 1010 nucleated cells containing 0.3% CD34+ progenitors. CD34+ cells were magnetically separated from the product to a final purity of 85% with a 64% yield. These results indicate that hematopoietic progenitors can safely be gathered from an MPSVII patient in numbers sufficient for the trial of clinical gene therapy applications. J. Clin. Apheresis 19:151–153, 2004. © 2004 Wiley‐Liss, Inc.
Assessment of complement activation during membrane‐based plasmapheresis procedures
Authors: Burnouf Thierry , Eber Michel , Kientz Daniel , Cazenave Jean‐Pierre , Burkhardt Thomas
Published: October 2004

Previous studies have suggested that plasmapheresis procedures using a separation membrane may activate the complement system and release anaphylatoxins. This study determines the content in C3a/C3ades Arg and C5a/C5ades Arg in plasma donations obtained by the new Haemonetics® Filter Core (FC) procedure and compares it to Baxter Autopheresis C® (Auto‐C). FC performs sequential blood centrifugation and plasma filtration on a microporous polyethersulfone membrane, while Auto‐C removes blood cells by simultaneous gravitation and filtration on a rotating nylon membrane. One group of 34 donors donated on FC and two groups of 30 and 10 donors on Auto‐C. Plasma aliquots were taken from the plasma units within 30 min of the end of the collection procedures, frozen at <−30°C and assessed for C3a and C5a at various time points of storage. Mean C3a/C3ades Arg in FC plasma (N = 34) was 1,151 (range: 526–2,991), 1,092 (range: 349–3498), and 507 (range: 307–815) ng/ml at time of collection and after 6 and 12 months of storage, respectively. Respective C5a/C5ades Arg was 26.6 (range 4.9–74), 18.9 (9.5–42.6), and 30.9 (range: 10.7–62.3) ng/ml. Mean C3a/C3ades Arg was higher in Auto‐C (P < 0.001): 4,724 ng/ml (N = 10; range: 2,400–7 ,360) and >4,149 ng/ml (N = 30; 2,408–>6,430) after 3 and 18 months storage, respectively. Mean C5a/C5ades Arg was 32.1 ng/ml (N = 30; range: 10.6–57.2) after 18 months of storage. Complement activation in FC plasmas appears limited compared to Auto‐C, suggesting better biocompatibility of this collection device and/or a favourable impact of the sequential cell centrifugation/filtration technology used. Further studies are needed to explain differences in complement activation between apheresis procedures and to assess clinical impacts, if any. J. Clin. Apheresis, 19:142–147, 2004. © 2004 Wiley‐Liss, Inc.
Plasmapheresis in acute disseminated encephalomyelitis
Authors: Lin Chin‐Hsien , Jeng Jiann‐Shing , Yip Ping‐Keung
Published: October 2004

High‐dose corticosteroid administration has been regarded as the main therapy for acute disseminated encephalomyelitis (ADEM). However, some patients with ADEM do not respond well to this treatment. We successfully used plasmapheresis to treat 2 patients who had ADEM. We also compared our patients' symptoms and clinical outcomes to those from previous reports. Plasmapheresis may be indicated not only for severe cases that fail to improve after high‐dose corticosteroid treatment but also for first‐line treatment. Additional large, controlled, double‐blinded trials are needed to clarify the role of plasmapheresis in ADEM. J Clin. Apheresis 19:154–159, 2004. © 2004 Wiley‐Liss, Inc.
Initiation of DNA replication at a nuclear matrix‐attached chromatin fraction
Authors: Radichev Ilian , Parashkevova Anastassia , Anachkova Boyka
Published: October 2004

It is still unclear what nuclear components support initiation of DNA replication. To address this issue, we developed a cell‐free replication system in which the nuclear matrix along with the residual matrix‐attached chromatin was used as a substrate for DNA replication. We found out that initiation occurred at late G1 residual chromatin but not at early G1 chromatin and depended on cytosolic and nuclear factors present in S phase cells but not in G1 cells. Initiation of DNA replication occurred at discrete replication foci in a pattern typical for early S phase. To prove that the observed initiation takes place at legitimate DNA replication origins, the in vitro synthesized nascent DNA strands were isolated and analyzed. It was shown that they were enriched in sequences from the core origin region of the early firing, dihydrofolate reductase origin of replication ori‐β and not in distal to the origin sequences. A conclusion is drawn that initiation of DNA replication occurs at discrete sub‐chromosomal structures attached to the nuclear matrix. © 2004 Wiley‐Liss, Inc.
JAK and STAT proteins are expressed and activated by IFN‐γ in rat pancreatic acinar cells
Authors: Gallmeier E. , Schäfer C. , Moubarak P. , Tietz A. , Plössl I. , Huss R. , Göke B. , Wagner A.C.C.
Published: October 2004

The development of acute pancreatitis (AP) is triggered by acinar events, but the subsequent extra‐acinar events, particularly a distinct immune response, appear to determine its severity. Cytokines modulate this immune response and are derived not only from immunocytes but also from pancreatic acinar cells. We studied whether pancreatic acinar cells were also capable of responding to cytokines. The JAK/STAT‐pathway represents the main effector for many cytokines. Therefore, expression and regulation of JAK and STAT proteins were investigated in rat pancreatic acinar cells. Western blotting showed expression of JAK1, JAK2, Tyk2, and STAT1, STAT2, STAT3, STAT5, STAT6. In addition, STAT1 was reversibly tyrosine‐phosphorylated upon the procedure of acinar cell isolation. In contrast, STAT3‐phosphorylation occurred spontaneously after pancreas removal and was not reversible within 8 h. STAT1 phosphorylation was also observed upon treatment with IFN‐γ but not upon EGF, TNF‐α or IL‐6, and inhibited by the JAK2‐inihibitor AG‐490. Immunohistochemistry revealed cytoplasmic expression of unphosphorylated STAT1 in untreated acinar cells and nuclear translocation of phosphorylated STAT1 following IFN‐γ‐treatment. Interestingly, although CCK leads to the activation of multiple stress pathways in pancreatic acinar cells, we found no influence of CCK on phosphorylation of STAT1, STAT3, or STAT5 in the pancreas. In conclusion, our data provide further evidence that pancreatic acinar cells are able to interact with immune cells. Besides stimulating immune cells via cytokine secretion, acinar cells are in turn capable of responding to IFN‐γ via JAK2 and STAT1 which may have an impact on the development of AP. © 2004 Wiley‐Liss, Inc.
Glioblastoma cells deficient in DNA‐dependent protein kinase are resistant to cell death
Authors: Chen George G. , Sin Fanny L.F. , Leung Billy C.S. , Ng Ho K. , Poon Wai S.
Published: October 2004

DNA‐dependent protein kinase (DNA‐PK), a nuclear serine/threonine kinase, is responsible for the DNA double‐strand break repair. Cells lacking or with dysfunctional DNA‐PK are often associated with mis‐repair, chromosome aberrations, and complex exchanges, all of which are known to contribute to the development of human cancers including glioblastoma. Two human glioblastoma cell lines were used in the experiment, M059J cells lacking the catalytic subunit of DNA‐PK, and their isogenic but DNA‐PK proficient counterpart, M059K. We found that M059K cells were much more sensitive to staurosporine (STS) treatment than M059J cells, as demonstrated by MTT assay, TUNEL detection, and annexin‐V and propidium iodide (PI) staining. A possible mechanism responsible for the different sensitivity in these two cell lines was explored by the examination of Bcl‐2, Bax, Bak, and Fas. The cell death stimulus increased anti‐apoptotic Bcl‐2 and decreased pro‐apoptotic Bcl‐2 members (Bak and Bax) and Fas in glioblastoma cells deficient in DNA‐PK. Activation of DNA‐PK is known to promote cell death of human tumor cells via modulation of p53, which can down‐regulate the anti‐apoptotic Bcl‐2 member proteins, induce pro‐apoptotic Bcl‐2 family members and promote a Bax–Bak interaction. Our experiment also demonstrated that the mode of glioblastoma cell death induced by STS consisted of both apoptosis and necrosis and the percentage of cell death in both modes was similar in glioblastoma cell lines either lacking DNA‐PK or containing intact DNA‐PK. Taken together, our findings suggest that DNA‐PK has a positive role in the regulation of apoptosis in human glioblastomas. The aberrant expression of Bcl‐2 family members and Fas was, at least in part, responsible for decreased sensitivity of DNA‐PK deficient glioblastoma cells to cell death stimuli. © 2004 Wiley‐Liss, Inc.
Specific AHNAK expression in brain endothelial cells with barrier properties
Authors: Gentil Benoît J. , Benaud Christelle , Delphin Christian , Remy Chantal , Berezowski Vincent , Cecchelli Roméo , Feraud Olivier , Vittet Daniel , Baudier Jacques
Published: October 2004

The blood–brain barrier (BBB) is essential for maintaining brain homeostasis and low permeability. Because disruption of the BBB may contribute to many brain disorders, they are of considerable interests in the identification of the molecular mechanisms of BBB development and integrity. We here report that the giant protein AHNAK is expressed at the plasma membrane of endothelial cells (ECs) forming specific blood–tissue barriers, but is absent from the endothelium of capillaries characterized by extensive molecular exchanges between blood and extracellular fluid. In the brain, AHNAK is widely distributed in ECs with BBB properties, where it co‐localizes with the tight junction protein ZO‐1. AHNAK is absent from the permeable brain ECs of the choroid plexus and is down‐regulated in permeable angiogenic ECs of brain tumors. In the choroid plexus, AHNAK accumulates at the tight junctions of the choroid epithelial cells that form the blood–cerebrospinal fluid (CSF) barrier. In EC cultures, the regulation of AHNAK expression and its localization corresponds to general criteria of a protein involved in barrier organization. AHNAK is up‐regulated by angiopoietin‐1 (Ang‐1), a morphogenic factor that regulates brain EC permeability. In bovine cerebral ECs co‐cultured with glial cells, AHNAK relocates from the cytosol to the plasma membrane when endothelial cells acquire BBB properties. Our results identify AHNAK as a protein marker of endothelial cells with barrier properties. © 2004 Wiley‐Liss, Inc.
Reorganization of ZO‐1 by sodium‐dependent glucose transporter activation after heat stress in LLC‐PK1 cells
Authors: Ikari Akira , Nakano Mika , Suketa Yasunobu , Harada Hitoshi , Takagi Kuniaki
Published: October 2004

Heat stress (HS) induces activation of high‐affinity sodium‐dependent glucose transporter (SGLT1) in porcine renal LLC‐PK1 cells. In this study, we investigated the roles of SGLT1 activation in reorganization of zonula occludens‐1 (ZO‐1), a cytosolic tight junction (TJ) protein, after HS. HS (42°C, 3 h) caused decrease in transepithelial electrical resistance (TER). Subsequent incubation at 37°C for 12 h increased TER above pre‐HS level. The treatment of phloridzin, a potent SGLT1 inhibitor, or the replacement of glucose with a nonmetabolizable glucose analog blocked the recovery of TER and increased the transepithelial flux of FITC‐dextran (4,000 Da). Immunofluorescent staining of ZO‐1 showed that HS diffused ZO‐1 from cell contact to cytosolic sites. Furthermore, the fraction of ZO‐1 was distributed from the Triton X‐100 insoluble to the Triton X‐100 soluble pool. After incubation at 37°C for 12 h, cell contact and ZO‐1 extractability with Triton X‐100 returned to pre‐HS conditions, but the recovery was completely prevented by phloridzin. Tyrosine kinases activity was increased by HS that was inhibited by phloridzin. Genistein and CGP77675, tyrosine kinases inhibitors, blocked the recovery of TER and increased the transepithelial flux of FITC‐dextran. Furthermore, these inhibitors prevented the recovery of cell contact and ZO‐1 extractability with Triton X‐100 as same as phloridzin. These findings suggested that the activation of SGLT1 reorganized ZO‐1 mediated by elevation of tyrosine kinases activity after heat injury. © 2004 Wiley‐Liss, Inc.
Heterogeneity of the immune response to adenovirus‐mediated factor VIII gene therapy in different inbred hemophilic mouse strains
Authors: Rawle Fiona E. M. , Shi Chang Xin , Brown Brian , McKinven Alexis , Tinlin Shawn , Graham Frank L. , Hough Christine , Lillicrap David
Published: October 2004

Background

The development of anti‐factor VIII (FVIII) antibodies (inhibitors) is a critical concern when considering gene therapy as a potential treatment modality for hemophilia A. We used a hemophilia A mouse model bred on different genetic backgrounds to explore genetically controlled differences in the immune response to FVIII gene therapy.

Methods

C57BL/6 FVIII knockout (C57‐FVIIIKO) mice were bred with normal BALB/c (BAL) mice, to generate a recombinant congenic BAL‐FVIIIKO model of hemophilia A. Early generation adenoviral (Ad) vectors containing the canine FVIII B‐domain‐deleted transgene under the control of either the CMV promoter or a tissue‐restricted (TR) promoter were administered to C57‐FVIIIKO, C57xBAL(F1)‐FVIIIKO crosses, and BAL‐FVIIIKO mice. FVIII expression, inhibitor development, inflammation, and vector‐mediated toxicity were assessed.

Results

In response to administration of Ad‐CMV‐cFVIII, C57‐FVIIIKO mice attain 3‐fold higher levels of FVIII expression than BAL‐FVIIIKO. All strains injected with Ad‐CMV‐FVIII displayed FVIII expression lasting only 2 weeks, with associated inhibitor development. C57‐FVIII‐KO mice that received Ad‐TR‐FVIII expressed FVIII for 12 months post‐injection, whereas FVIII expression was limited to 1 week in C57xBAL(F1)‐FVIIIKO and BAL‐FVIIIKO mice. This loss of expression was associated with anti‐FVIII inhibitor development. BAL‐FVIIIKO mice showed increased hepatotoxicity with alanine aminotransferase levels reaching 4‐fold higher levels than C57‐FVIIIKO mice. However, C57‐FVIIIKO mice initiate a more rapid and effective cell‐mediated clearance of virally transduced cells than BAL‐FVIIIKO, as evidenced by real‐time PCR analysis of transduced tissues. Overall, strain‐dependent differences in the immune response to FVIII gene delivery were only noted in the adaptive response, and not in the innate response.

Conclusions

Our results indicate that the genetic background of the murine model of hemophilia A influences FVIII expression levels, the development of anti‐FVIII inhibitors, clearance of transduced cells, and the severity of vector‐mediated hepatotoxicity. Copyright © 2004 John Wiley & Sons, Ltd.
Purine nucleoside phosphorylase and fludarabine phosphate gene‐directed enzyme prodrug therapy suppresses primary tumour growth and pseudo‐metastases in a mouse model of prostate cancer
Authors: Martiniello‐Wilks Rosetta , Wang Xiao Yang , Voeks Dale J. , Dane Allison , Shaw Jan M. , Mortensen Elin , Both Gerald W. , Russell Pamela J.
Published: October 2004

Gene‐directed enzyme prodrug therapy based on the E. coli purine nucleoside phosphorylase (PNP) gene produces efficient tumour cell killing. PNP converts adenosine analogs into toxic metabolites that diffuse across cell membranes to kill neighbouring untransduced cells (PNP‐GDEPT). Interference with DNA, RNA and protein synthesis kills dividing and non‐dividing cells, an important consideration for slow‐growing prostate tumours. This study examined the impact of administering PNP‐GDEPT into orthotopically grown RM1 prostate cancers (PCas) on the growth of lung pseudo‐metastases of immunocompetent mice. C57BL/6 mice bearing orthotopic RM1 PCas received a single intraprostatic injection of OAdV220 (1010 particles), a recombinant ovine atadenovirus containing the PNP gene controlled by the Rous Sarcoma virus promoter, followed by fludarabine phosphate (∼600 mg/m2/day) administered intraperitoneally (ip) once daily for 5 days. Pseudo‐metastases were induced 2 days after intraprostatic vector administration by tail‐vein injection of untransduced RM1 cells. Mice given PNP‐GDEPT showed a significant reduction both in prostate volume (∼50%) and in lung colony counts (∼60%). Apoptosis was increased two‐fold in GDEPT‐treated prostates compared with controls (P < 0.01), but was absent in the lungs. Staining for proliferating cell nuclear antigen (PCNA) indicated that proliferation of both RM1 prostate tumours (P < 0.01) and lung colonies (P < 0.01) was significantly suppressed after GDEPT. Although prostate tumour immune cell infiltration did not differ significantly between treatments, immunostaining for Thy‐1.2 (CD90) showed that GDEPT promoted Thy‐1.2+ cell infiltration into the prostate tumour site. This study showed that a single course of PNP‐GDEPT significantly suppressed local PCa growth and reduced lung colony formation in the aggressive RM1 tumour model. Copyright © 2004 John Wiley & Sons, Ltd.
Inter‐patient variation in efficacy of five oncolytic adenovirus candidates for ovarian cancer therapy
Authors: Lam John T. , Kanerva Anna , Bauerschmitz Gerd J. , Takayama Koichi , Suzuki Kaori , Yamamoto Masato , Bhoola Snehal M. , Liu Bin , Wang Minghui , Barnes Mack N. , Alvarez Ronald D. , Siegal Gene P. , Curiel David T. , Hemminki Akseli
Published: October 2004

Background

Gene therapy offers a new strategy for cancer treatment. Adenoviruses represent the most widely used gene therapy vector and feature an excellent safety record. Conditionally replicative adenoviruses (CRAds) effect solid tumor penetration and tumor selective oncolysis and consequently offer potential efficacy for metastatic disease treatment. We evaluated five CRAds as candidate clinical agents for ovarian cancer therapy: RGDCRADcox‐2R, Ad5VEGFE1, Ad5/3VEGFE1, Ad5‐Δ24RGD, and Ad5/3‐Δ24.

Methods

DNA replication by these five CRAds, wild‐type adenovirus, and an E1‐deleted control was measured in purified primary ovarian cancer cell spheroids by quantitative PCR. CRAd‐mediated oncolysis was quantified in ovarian cancer cell monolayers and three‐dimensional spheroids by cellular viability assays. The therapeutic efficacy of each CRAd was tested by intraperitoneal administration in mice with peritoneally disseminated human ovarian cancer.

Results

An increase in viral DNA was noted in primary tumor cell spheroids for all replicative viruses tested. Variation was noted in viral DNA replication between patient samples. All five CRAds induced remarkable oncolysis. They also prolonged survival in vivo compared with the wild‐type control group.

Conclusions

All five CRAds tested showed robust DNA replication, oncolysis, and in vivo therapeutic efficacy. Each virus has potential for clinical testing, and such further testing will ultimately determine its safety and relative usefulness. Variation of CRAd DNA replication between different patient samples suggests that target tissue features, such as surface receptors and endogenous transcription factors, may affect CRAd infectivity and replicativity. Evaluation of such factors may become important to optimize cancer therapy for individual patients. Copyright © 2004 John Wiley & Sons, Ltd.
Efficient hybrid scheme of finite element method of lines for modelling computational electromagnetic problems
Authors: Chen R. S. , Yung Edward K. N. , Wu Ke
Published: October 2004

A hybrid scheme called finite element method of lines is proposed and described for modelling and analysis of generalized computational electromagnetic problems with emphasis on a number of irregular waveguide examples. This new technique is developed by combining a finite element method with a method of lines so that it can handle not only irregular composite geometry but also maintain high accuracy enjoyed by semi‐analytical procedures. Analytical and numerical algorithmic building blocks of this new scheme are discussed in detail such as geometry discretization, element mapping, element trial functions, reformulation and computational issues of non‐linear ordinary differential equations. Our results show that this new technique is able to efficiently solve complex problems as compared with the conventional method of lines. Copyright © 2004 John Wiley & Sons, Ltd.
Fungicidal effect of diode laser irradiation in patients with denture stomatitis
Authors: Maver‐Biscanin Mirela , Mravak‐Stipetic Marinka , Jerolimov Vjekoslav , Biscanin Alen
Published: October 2004

Background and Objective

Denture stomatitis (DS) is a common inflammatory condition that affects denture wearers. The aim of this study was to examine, in vivo, the effect of diode laser irradiation on fungal growth in both the palatal mucosa and in denture base materials, in denture wearing patients.

Study Design/Materials and Methods

In total, 70 patients with clinical study design evidence of DS participated in this parallel, single blind, and placebo controlled study. The subjects were randomly assigned to one of four different treatment regimens: (1) irradiation with a 685 nm wavelength laser for 10 minutes (30 mW); (2) irradiation with a 830 nm wavelength laser for 5 minutes (60 mW). A semiconductor diode laser, BTL‐2000 (BTL‐2 Dravotnicka Technika, Prague, Czech Republic), was used in both treatment cases using an energy density of 3.0 J/cm2 and a continuous working mode for five consecutive days; (3) placebo—sham irradiation of patients; (4) antimicotic—self treatment of patient's palatal mucosa with an antifungal oral gel and the use of an antiseptic solution for their dentures. The effect of laser light on fungal growth in vivo was evaluated after final treatment using the swab method and a semi‐quantitative estimation of Candida albicans colonies cultivated on agar plates.

Results

A fungicidal effect was achieved in the laser treated and antimicotic treated groups, whereas most subjects in the placebo group were found to have unchanged conditions on both their palate (P = 0,004) and dentures (P < 0,001).

Conclusions

Light from a low‐power laser (LLLT) may be valuable in the treatment of DS. This is of great importance since the rate of recurrence of disease is high, whereas an optimal treatment modality has not yet been found. Lasers Surg. Med. 35:259–262, 2004. © 2004 Wiley‐Liss, Inc.
Ultrastructural comparison of medial collateral ligament repair after single or multiple applications of GaAlAs laser in rats
Authors: Ng Gabriel Y.F. , Fung Dicky T.C. , Leung Mason C.P. , Guo Xia
Published: October 2004

Background and Objectives

To examine single versus multiple applications of a gallium aluminum arsenide (GaAlAs) laser on the ultrastructural morphology of surgically injured medial collateral ligaments (MCLs) in rats.

Study Design/Materials and Methods

Sixteen rats were studied with 12 receiving right MCL transection and 4 receiving sham injury. Group 1 (n = 4) received one session of laser (31.6 J/cm2) immediately after injury. Group 2 (n = 4) received 9 doses of transcutaneous laser (3.5 J/cm2). The controls (Group 3, n = 4) received one session of placebo laser, while the sham Group 4 (n = 4) received no treatment. Ultrastructural analyses were done with electron microscopy at 3 weeks.

Results

The mass‐averaged diameters of collagen fibril in the core and periphery of MCLs treated with multiple laser were larger than the control and those with single laser treatment (P < 0.05). However, the sham injured group had larger fibrils than all other groups (P < 0.05).

Conclusions

The repairing MCLs had smaller collagen fibrils than the sham injured ligaments. Multiple laser treatments enhanced the collagen growth in the repairing MCLs at 3 weeks after injury, which are superior to a single treatment with similar dosage. Lasers Surg. Med. 35:317–323, 2004. © 2004 Wiley‐Liss, Inc.
Enhancement of the immune response to residual intrahepatic tumor tissue by laser‐induced thermotherapy (LITT) compared to hepatic resection
Authors: Isbert Christoph , Ritz Jörg‐P. , Roggan André , Schuppan Detlef , Rühl Martin , Buhr Heinz J. , Germer Christoph‐T.
Published: October 2004

Background and Objectives

In contrast to hepatic resection, thermally destroyed autologous tumor cells remain in situ after laser‐induced thermotherapy (LITT). The aim of the study was to evaluate the effect of LITT and hepatic resection on the immune response to residual intrahepatic tumor tissue and the growth of untreated liver metastases.

Study Design/Materials and Methods

Two independent adenocarcinomas (CC531) were implanted into 60 WAG rats, one in the right (control tumor) and one in the left liver lobe (treated tumor). The left lobe tumor was treated either by LITT or partial hepatectomy. The control tumor was submitted to further investigation 24 hours, 96 hours, 7 days, and 10 days after treatment.

Results

Ten days after treatment, control tumor volumes were 296±46 mm_ after LITT and 1,181±192 mm_, 1,387±200 mm_ after hepatic resection and no treatment, respectively (P<0.001). Peritoneal tumor spread was detected in 4/20 cases after LITT and in 17/20 cases after hepatic resection. Expression of CD8, B7‐2 (CD86), and to lesser extent MHCII, LFA1 (CD11a), and ICAM1 (CD54), was significantly enhanced at the invasion front of control tumors after LITT compared to hepatic resection.

Conclusions

Our results suggest that LITT increases the immune response against untreated intrahepatic tumor tissue, which can lead to reduced tumor growth. Lasers Surg. Med. 35:284–292, 2004. © 2004 Wiley‐Liss, Inc.
Selective targeting of protein, water, and mineral in dentin using UV and IR pulse lasers: The effect on the bond strength to composite restorative materials
Authors: Sheth Karishma K. , Staninec Michal , Sarma Anupama V. , Fried Daniel
Published: October 2004

Background and Objectives

Previous studies have shown that during the laser irradiation of dentin and bone, thermal damage can be minimized by using a highly absorbed laser wavelength, laser pulses shorter than the thermal relaxation time of the deposited laser energy at that wavelength, and the addition of a layer of water to the tissue surface before ablation. The objective of this study was to investigate the influence of laser pulse duration and wavelength with and without the added water layer on the bond strength of composite to laser prepared dentin surfaces. The specific hypothesis that was tested was that thermal damage to the collagen matrix in dentin compromises the bond strength to composite restorative materials.

Study Design/Materials and Methods

Three laser systems were employed that were tuned to water, collagen, and mineral absorption with pulse durations less than the thermal relaxation time of the deposited energy. The surfaces of human dentin were irradiated by laser irradiation from free‐running and Q‐switched Er:YSGG lasers, pulsed CO2 lasers operating at 9.6‐μm, and a Q‐switched Nd:YAG laser operating at 355‐nm. A motion control system and a pressurized spray system incorporating a microprocessor controlled pulsed nozzle for water delivery, were used to ensure uniform treatment of the entire surface. Shear bond testing was used to evaluate the adhesive strength in order to access the suitability of laser treated surfaces for bonding. Bonded interfaces were examined by SEM.

Results

All the laser groups had significantly lower bond strengths than the positive acid etch control group. The highest bond strengths were for the short pulse (< 5‐microsecond) Er:YSGG and CO2 laser groups with water. Laser groups without water had significantly reduced bond strengths and thicker layers of thermally damaged dentin.

Conclusions

Thermal damage to the collagen matrix profoundly influences the bond strength to composite restorations. Lasers Surg. Med. 35:245–253, 2004. © 2004 Wiley‐Liss, Inc.
Dental bleaching efficacy with diode laser and LED irradiation: An in vitro study
Authors: Wetter Niklaus Ursus , Barroso Márcia C.S. , Pelino José E.P.
Published: October 2004

Background and Objectives

Bleaching techniques achieved significant advances with the use of coherent or incoherent radiation sources to activate the bleaching agents. This in vitro study compares the whitening efficacy of LED and diode laser irradiation using the two agents Opalescence X‐tra and HP Whiteness.

Study Design/Materials and Methods

A total of 60 bovine incisors were randomly divided into six groups, three for each bleaching agent, receiving only agent, agent and LED (wavelength 470 nm), agent and 1.6 W diode laser (808 nm). The results of the irradiations were characterized using the CIEL*a*b* system.

Results

Significant differences in the chroma value are obtained for the two whitening agents and for the different light sources. In terms of lightness, the association of Laser and Whiteness HP bleaching gel showed significantly better results than when the same agent was used alone or in combination with LED.

Conclusions

Best overall results are obtained with the Whiteness HP and Laser association. Lasers Surg. Med. 35:254–258, 2004. © 2004 Wiley‐Liss, Inc.
Self‐Similar Wave of Swelling/Collapse Phase Transition along Polyelectrolyte Gel
Authors: Zubarev Andrey Yu. , Blyakhman Felix A. , Pollack Gerald H. , Gusev Peter , Safronov Alexander P.
Published: October 2004

Summary: Theoretical analysis of the possibility of collapse/swelling phase transition propagation along a polyelectrolyte gel thread has been performed. A differential equation that determines the time dependence of the degree of swelling of polymer thread under the radial mechanical force has been obtained. It formally coincides with the equation of diffusion of an impurity in a medium with a certain source density that depends on excluded‐volume and Coulomb interactions, osmotic pressure of counterions, and entropy of the subchains. Two stationary points of this equation correspond to contracted and swollen states. It has been shown that once activated, the phase transition between these stationary points could propagate along the thread in the form of a wave with positive speed if the initial state is characterized by a higher level of free energy than the final state. The results of numerical calculations illustrating the process of phase transition propagation along polyelectrolyte gel thread show that propagation of the swelling/collapse phase transition takes place in the form of the stepwise self‐similar wave with steep front. The sharp boundary between swollen and collapsed zones is observed and does not tend to dissipate as propagation proceeds.

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Interfacial Thermodynamics of Polymeric Mesophases
Authors: Rey Alejandro D.
Published: October 2004

Summary:A complete mechanical‐thermodynamical formulation for multicomponent nematic polymer‐isotropic fluid interfaces is derived, validated, and used to derive the structure and shape equations for these soft anisotropic polymer interfaces. The fundamental role of liquid crystalline order and long range effects in coupling bulk and interfacial effects, and in coupling thermodynamical/liquid crystalline order/geometrical variables is demonstrated, discussed, and validated. The Gibbs‐Duhem nemato‐thermodynamics equation emerges from an interfacial tension γ = γ(Θ, μ,Q, ∇sQ,k) that depends on temperature (Θ), chemical potential (μ), nematic tensor order parameterQ, surface gradients ofQ, and geometryk, and leads to new couplings in these enhanced phase spaces. The role of entropy and adsorption, and long range effects on interfacial shape and structure selection is revealed. For flat interfaces the preferred structure emerges from a competition between energy, entropy, and adsorption.
Chain Conformations in the Crystalline Field of Syndiotactic Vinyl Polymers Deriving from 1,3‐Diene Monomers. Analysis by Molecular Mechanics
Authors: Pirozzi Beniamino , Napolitano Roberto , Esposito Simona
Published: October 2004

Summary: Conformational energy calculations on the chain conformation in the crystalline field have been performed for various syndiotactic vinyl polymers deriving from 1,3‐diene monomers. Energy maps as a function of the independent torsion angles have evidenced for all the polymers minima corresponding to highly extended and to helical chains. Energy minimizations as a function of all the internal parameters for the s(2/1)2 and tcm symmetries have allowed the evaluation of the energy differences between chains having the two symmetries and the prediction of the values of the conformational parameters for each polymer. The results have been compared with the experimental data reported in the literature for some of the studied polymers.

Conformational energy map of sPBD12.

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Conformational energy map of sPBD12.
Validation of thea posteriorierror estimator based on polynomial preserving recovery for linear elements
Authors: Zhang Zhimin , Naga Ahmed
Published: October 2004

The purpose of this work is to investigate the quality of thea posteriorierror estimator based on the polynomial preserving recovery (PPR). The main tool in this investigation is the computer‐based theory. Also, a comparison is made between this estimator and the one based on the superconvergence patch recovery (SPR). The results of this comparison were found to be in favour of the estimator based on the PPR. Copyright © 2004 John Wiley & Sons, Ltd.
Bounds for element size in a variable stiffness cohesive finite element model
Authors: Tomar Vikas , Zhai Jun , Zhou Min
Published: October 2004

The cohesive finite element method (CFEM) allows explicit modelling of fracture processes. One form of CFEM models integrates cohesive surfaces along all finite element boundaries, facilitating the explicit resolution of arbitrary fracture paths and fracture patterns. This framework also permits explicit account of arbitrary microstructures with multiple length scales, allowing the effects of material heterogeneity, phase morphology, phase size and phase distribution to be quantified. However, use of this form of CFEM with cohesive traction–separation laws with finite initial stiffness imposes two competing requirements on the finite element size. On one hand, an upper bound is needed to ensure that fields within crack‐tip cohesive zones are accurately described. On the other hand, a lower bound is also required to ensure that the discrete model closely approximates the physical problem at hand. Both issues are analysed in this paper within the context of fracture in multi‐phase composite microstructures and a variable stiffness bilinear cohesive model. The resulting criterion for solution convergence is given for meshes with uniform, cross‐triangle elements. A series of calculations is carried out to illustrate the issues discussed and to verify the criterion given. These simulations concern dynamic crack growth in an Al2O3 ceramic and in an Al2O3/TiB2 ceramic composite whose phases are modelled as being hyperelastic in constitutive behaviour. Copyright © 2004 John Wiley & Sons, Ltd.
Conforming polygonal finite elements
Authors: Sukumar N. , Tabarraei A.
Published: October 2004

In this paper, conforming finite elements on polygon meshes are developed. Polygonal finite elements provide greater flexibility in mesh generation and are better‐suited for applications in solid mechanics which involve a significant change in the topology of the material domain. In this study, recent advances in meshfree approximations, computational geometry, and computer graphics are used to construct different trial and test approximations on polygonal elements. A particular and notable contribution is the use of meshfree (natural‐neighbour, nn) basis functions on a canonical element combined with an affine map to construct conforming approximations on convex polygons. This numerical formulation enables the construction of conforming approximation on n‐gons (n⩾3), and hence extends the potential applications of finite elements to convex polygons of arbitrary order. Numerical experiments on second‐order elliptic boundary‐value problems are presented to demonstrate the accuracy and convergence of the proposed method. Copyright © 2004 John Wiley & Sons, Ltd.