bluuebunny/crossref_metadata_2025 huggingface dataset:
PGE2 receptor EP1 renders dopaminergic neurons selectively vulnerable to low‐level oxidative stress and direct PGE2 neurotoxicity
Authors: Carrasco Emilce , Casper Diana , Werner Peter
Published: September 2007

Oxidative stress and increased cyclooxygenase‐2 (COX‐2) activity are both implicated in the loss of dopaminergic neurons from the substantia nigra (SN) in idiopathic Parkinson's disease (PD). Prostaglandin E2 (PGE2) is one of the key products of COX‐2 activity and PGE2 production is increased in PD. However, little is known about its role in the selective death of dopaminergic neurons. Previously, we showed that oxidative stress evoked by low concentrations of 6‐hydroxydopamine (6‐OHDA) was selective for dopaminergic neurons in culture and fully dependent on COX‐2 activity. We postulated that this loss was mediated by PGE2 acting through its receptors, EP1, EP2, EP3, and EP4. Using double‐label immunohistochemistry for specific EP receptors and tyrosine hydroxylase (TH), we identified EP1 and EP2 receptors on dopaminergic neurons in rat SN. EP2 receptors were also found in non‐dopaminergic neurons of this nucleus, as were EP3 receptors, whereas the EP4 receptor was absent. PGE2, 16‐phenyl tetranor PGE2 (a stable synthetic analogue), and 17‐phenyl trinor PGE2 (an EP1 receptor–selective agonist) were significantly toxic to dopaminergic cells at nanomolar concentrations; EP2‐ and EP3‐selective agonists were not. We challenged dopaminergic neurons in embryonic rat mesencephalic primary neuronal cultures and tested whether these receptors mediate selective 6‐OHDA toxicity. The nonselective EP1–3 receptor antagonist AH‐6809 and two selective EP1 antagonists, SC‐19220 and SC‐51089, completely prevented the 40%–50% loss of dopaminergic neurons caused by exposure to 5 μM 6‐OHDA. Together, these results strongly implicate PGE2 activation of EP1 receptors as a mediator of selective toxicity in this model of dopaminergic cell loss. © 2007 Wiley‐Liss, Inc.
Novel culture strategy for human stem cell proliferation and neuronal differentiation
Authors: Serra Margarida , Leite Sofia B. , Brito Catarina , Costa Júlia , Carrondo Manuel J.T. , Alves Paula M.
Published: September 2007

Embryonal carcinoma (EC) stem cells derived from germ cell tumors closely resemble embryonic stem (ES) cells and are valuable tools for the study of embryogenesis. Human pluripotent NT2 cell line, derived from a teratocarcinoma, can be induced to differentiate into neurons (NT2‐N) after retinoic acid treatment. To realize the full potential of stem cells, developing in vitro methods for stem cell proliferation and differentiation is a key challenge. Herein, a novel culture strategy for NT2 neuronal differentiation was developed to expand NT2‐N neurons, reduce the time required for the differentiation process, and increase the final yields of NT2‐N neurons. NT2 cells were cultured as 3D cell aggregates (“neurospheres”) in the presence of retinoic acid, using small‐scale stirred bioreactors; it was possible to obtain a homogeneous neurosphere population, which can be transferred for further neuronal selection onto coated surfaces. This culturing strategy yields higher amounts of NT2‐N neurons with increased purity compared with the amounts routinely obtained with static cultures. Moreover, mechanical and enzymatic methods for neurosphere dissociation were evaluated for their ability to recover neurons, trypsin digestion yielding the best results. Nevertheless, the highest recoveries were obtained when neurospheres were collected directly to treated surfaces without dissociation steps. This novel culture strategy allows drastic improvement in the neuronal differentiation efficiency of NT2 cells, insofar as a fourfold increase was obtained, reducing simultaneously the time needed for the differentiation process. The culture method described herein ensures efficient, reproducible, and scaleable ES cell proliferation and differentiation, contributing to the usefulness of stem cell bioengineering. © 2007 Wiley‐Liss, Inc.
Cellular and molecular characterization of oxidative stress in olfactory epithelium of Harlequin mutant mouse
Authors: Vaishnav Radhika A. , Getchell Marilyn L. , Huang Liping , Hersh Matthew A. , Stromberg Arnold J. , Getchell Thomas V.
Published: September 2007

Oxidative stress in the olfactory system is a major factor associated with age‐related olfactory impairment, although the mechanisms by which this occurs are not completely understood. The Harlequin mutant mouse (Hq/Y), which carries an X‐linked recessive mutation in the Aifm1 gene, is a model of progressive oxidative stress–induced neurodegeneration in the cerebellum and retina. To determine whether the Hq/Y mutant mouse is a suitable model of oxidative stress–associated olfactory aging, we investigated cellular and molecular changes in the olfactory epithelium (OE) and olfactory bulb (OB) of 6‐month‐old male Hq/Y mice compared to those in sex‐matched littermate controls (+/Y) and in age‐ and sex‐matched C57BL/6 mice. Immunoreactivity for apoptosis‐inducing factor, the protein product of Aifm1, was localized in mature olfactory sensory neurons (mOSNs) in +/Y mice but was rarely detected in Hq/Y mice. Hq/Y mice also exhibited increased lipofuscin autofluorescence and increased immunoreactivity for an oxidative DNA/RNA damage marker in mOSNs and in mitral/tufted cells in the OB and an increased number of cleaved caspase‐3 immunoreactive apoptotic cells in the OE. Microarray analysis demonstrated that Aifm1 expression was down‐regulated by 80% in the OE of Hq/Y mice compared to that in +/Y mice. Most significantly, regulated genes were classified into functional categories of cell signaling/apoptosis/cell cycle, oxidative stress/aging, and cytoskeleton/extracellular matrix/transport‐associated. Analysis with EASE software indicated that the functional categories significantly overrepresented in Hq/Y mice included up‐regulated mitochondrial genes and down‐regulated cytoskeletal organization‐ and neurogenesis‐related genes. Our results strongly support the Hq/Y mutant mouse being a novel model for mechanistic studies of oxidative stress–associated olfactory aging. © 2007 Wiley‐Liss, Inc.
Inhibition of proteoglycan synthesis affects neuronal outgrowth and astrocytic migration in organotypic cultures of fetal ventral mesencephalon
Authors: Berglöf Elisabet , Plantman Stefan , Johansson Saga , Strömberg Ingrid
Published: September 2007

Grafting fetal ventral mesencephalon has been utilized to alleviate the symptoms of Parkinson's disease. One obstacle in using this approach is the limited outgrowth from the transplanted dopamine neurons. Thus, it is important to evaluate factors that promote outgrowth from fetal dopamine neurons. Proteoglycans (PGs) are extracellular matrix molecules that modulate neuritic growth. This study was performed to evaluate the role of PGs in dopamine nerve fiber formation in organotypic slice cultures of fetal ventral mesencephalon. Cultures were treated with the PG synthesis inhibitor methyl‐umbelliferyl‐β‐D‐xyloside (β‐xyloside) and analyzed using antibodies against tyrosine hydroxylase (TH) to visualize dopamine neurons, S100β to visualize astrocytes, and neurocan to detect PGs. Two growth patterns of TH‐positive outgrowth were observed: nerve fibers formed in the presence of astrocytes and nerve fibers formed in the absence of astrocytes. Treatment with β‐xyloside significantly reduced the distance of glial‐associated TH‐positive nerve fiber outgrowth but did not affect the length of the non‐glial‐associated nerve fibers. The addition of β‐xyloside shifted the nerve fiber growth pattern from being mostly glial‐guided to being non‐glial‐associated, whereas the total amount of TH protein was not affected. Further, astrocytic migration and proliferation were impaired after β‐xyloside treatment, and levels of non‐intact PG increased. β‐Xyloside treatment changed the distribution of neurocan in astrocytes, from being localized in vesicles to being diffusely immunoreactive in the processes. To conclude, inhibition of PG synthesis affects glial‐associated TH‐positive nerve fiber formation in ventral mesencephalic cultures, which might be an indirect effect of impaired astrocytic migration. © 2007 Wiley‐Liss, Inc.
Mode of cembranoid action on embryonic muscle acetylcholine receptor
Authors: Ulrich H. , Akk G. , Nery A.A. , Trujillo C.A. , Rodriguez A.D. , Eterović V.A.
Published: September 2007

The mechanism of eupalmerin acetate (EUAC) actions on the embryonic muscle nicotinic acetylcholine receptor (nAChR) in BC3H‐1 cells was studied by using whole‐cell and single‐channel patch‐clamp current measurements. With whole‐cell currents, EUAC did not act as an agonist on this receptor. Coapplication of 30 μM EUAC with 50 μM, 100 μM, or 500 μM carbamoylcholine (CCh) reversibly inhibited the current amplitude, whereas, with 20 μM CCh, current was increased above control values in the presence of EUAC. EUAC concentration curves (0.01–40 μM) obtained with 100 μM and 500 μM CCh displayed slope coefficients, nH, significantly smaller than one, suggesting that EUAC bound to several sites with widely differing affinities on the receptor molecule. The apparent rate of receptor desensitization in the presence of EUAC and CCh was either slower than or equal to that obtained with CCh alone. The major finding from single‐channel studies was that EUAC did not affect single‐channel conductance or the ability of CCh to interact with the receptor. Instead, EUAC acted by increasing the channel closing rate constant. The results are not consistent with the competitive model for EUAC inhibition, with the sequential open‐channel block model, or with inhibition by increased desensitization. The data are best accounted for by a model in which EUAC acts by closed‐channel block at low concentrations, by positive modulation at intermediate concentrations, and by negative allosteric modulation of the open channel at high concentrations. © 2007 Wiley‐Liss, Inc.
Histological correlation of diffusion tensor imaging metrics in experimental spinal cord injury
Authors: Herrera Juan J. , Chacko Tessy , Narayana Ponnada A.
Published: September 2007

Diffusion tensor imaging (DTI) has the potential to provide important information about the integrity of white matter tracts in injured spinal cord tissue. It is thought that DTI‐based transverse diffusivity (λt) reflects the state of myelin, whereas longitudinal diffusivity (λl) reflects axonal integrity. However, this has not been established in spinal cord injury (SCI). Therefore, we performed quantitative histologic analysis on 4‐ and 8‐week post‐SCI rodent spinal cords that had received a moderately severe injury at the T7 level and correlated the histology with λt and λl measured in vivo. Using antibodies specific to myelin and axonal process (i.e., neurofilament), the percent area of expression was determined in the dorsal, ventral, and lateral white matter from both rostral and caudal regions away from the epicenter of the injury site. The results suggest a positive correlation between λt and demyelination in many but not all regions. However, these studies failed to establish a correlation between λl and axonal damage. These results suggest that caution must be exercised in interpreting the DTI metrics in terms of tissue pathology in SCI. © 2007 Wiley‐Liss, Inc.
Proteomic analysis of embryonic stem cell–derived neural cells exposed to the antidepressant paroxetine
Authors: McHugh Patrick C. , Rogers Geraldine R. , Loudon Barbara , Glubb Dylan M. , Joyce Peter R. , Kennedy Martin A.
Published: September 2007

Antidepressant drugs can have significant effects on the mood of a patient suffering from major depression or other disorders. The pharmacological actions of these drugs generally affect the uptake or metabolism of the neurotransmitters serotonin, noradrenalin, and, to a lesser extent, dopamine. However, many aspects of antidepressant action are not understood. We conducted a proteomic analysis in a neuronal cell culture model in an attempt to identify molecules important to the operation of pathways functionally relevant to antidepressant action. The model involved generating cultures containing mixed neural and glial cells by controlled differentiation of mouse embryonic stem cells, followed by exposure to 1 μM paroxetine for 14 days. After antidepressant exposure, we observed increased expression or modification of sepiapterin reductase (SPR), heat shock protein 9A, RAS and EF‐hand domain containing, and protein disulfide isomerase associated 3 and decreased expression or modification of creatine kinase, actin, prohibitin, a T‐cell receptor α chain, defensin‐related cryptdin 5, and the intermediate filament proteins glial fibrillary acidic protein and vimentin. SPR, the most strongly up‐regulated protein observed, controls production of tetrahydrobiopterin, an essential cofactor for the synthesis of many neurotransmitters including serotonin, making it a plausible and intriguing candidate protein for involvement in mood control and antidepressant drug action. SPR and the other proteins identified may represent links to molecular processes of importance to mood dysregulation and control, and their respective genes may be novel candidates for the study of antidepressant pharmacogenetics. © 2007 Wiley‐Liss, Inc.
The use of integra artificial dermis to minimize donor‐site morbidity after suprafascial dissection of the radial forearm flap
Authors: Gravvanis Andreas I. , Tsoutsos Dimosthenis A. , Iconomou Thomais , Gremoutis George
Published: September 2007

In an effort to minimize the radial forearm flap donor‐site morbidity, the flap was elevated using the suprafascial dissection technique, in six patients with various facial defects. The donor site was covered primarily with Integra artificial skin and secondarily with an ultrathin split‐thickness skin graft. The mean time to wound healing of the forearm donor site was 24 days. There were no flap failures, and all flaps healed uneventfully. At the end of the follow‐up, all patients showed normal range of motion of the wrist and the fingers, normal power grip, and power pinch. All patients evaluated the esthetic appearance of the forearm donor site as very good. In conclusion, suprafascial dissection of the forearm flap creates a superior graft recipient site, and the use of Integra artificial dermis is a valuable advancement to further minimize the donor‐site morbidity, resulting in excellent functional and aesthetic outcomes. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
Selection of recipient vessels in microsurgical free tissue reconstruction of head and neck defects
Authors: Yazar Sukru
Published: September 2007

The development of microsurgical techniques has facilitated proper management of extensive head and neck defects and deformities. Bone or soft tissue can be selected to permit reconstruction with functional and aesthetic results. However, for free tissue transfer to be successful, proper selection of receipient vessels is as essential as the many other factors that affect the final result. In this article selection strategies for recipient vessels for osteocutaneous free flaps, soft tissue free flaps, previously dissected and irradiated areas, recurrent and subsequent secondary reconstructions, simultaneous double free flap transfers in reconstruction of extensive composite head and neck defects, and the selection of recipient veins are reviewed in order to provide an algorithm for the selection of recipient vessels for head and neck reconstruction. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
Reconstruction of the human median nerve in the forearm with the Neurotube™
Authors: Donoghoe Nicholas , Rosson Gedge D. , Dellon A. Lee
Published: September 2007

Reconstruction of the median nerve in the forearm traditionally has required multiple interfascicular interposition autologous nerve grafts. Recovery of two‐point discrimination, even in young adults, has been reported rarely. Two patients, aged 43 and 61, are reported with 5‐year follow‐up of median nerve reconstruction in the distal forearm in which multiple interposition bioabsorbable polyglycolic acid conduits (Neurotube™) were used to bridge the 3‐cm nerve defects. Four separate 2.3‐mm diameter, 4.0‐cm long conduits were used in each patient. Sensory re‐education was used for rehabilitation. Each patient recovered two‐point discrimination with good localization in the thumb, index, and middle finger by 2 years after the nerve reconstruction. Both patients recovered abductor pollicis brevis function. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
Surgical treatment of superimposed nerve compressions in hepatitis C neuropathy
Authors: Rosson Gedge D. , Rodriguez Eduardo D. , Dellon A. Lee
Published: September 2007

The plastic surgeon's usual involvement in patients with hepatitis C is most frequently limited to an inner city population with hand and forearm abscesses from intravenous drug use or to incidences of needle‐stick injury in the operating room when the patient is hepatitis C positive. Hand surgeons and peripheral nerve surgeons often treat patients with underlying neuropathies who have superimposed overlying nerve compressions such as carpal tunnel syndrome. We have applied this experience to a patient with underlying peripheral neuropathy associated with Hepatitis C and clinical evidence of overlying lower extremity nerve compressions. We believe that she is the first successful surgical treatment of peripheral nerve compressions in a patient with hepatitis C—associated neuropathy, documented by noninvasive neurosensory testing. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
Clinical experience with the nasolabial fold as receptor site in microvascular reconstruction
Authors: Smit Jeroen M. , Hartman ED H. M. , Acosta Rafael
Published: September 2007

We report our experience using the vessels at the nasolabial fold as receptor site in free tissue transfer in head and neck reconstructions; a site that proved more convenient than the submandibulary site in selected cases. Six cases as well as the dissection technique of the nasolabial fold are reported. No complications occurred during or post surgery and in all cases the vessels were of adequate diameter for an end to end anastomosis. The advantages this site offers are discussed. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
Early reintervention of compromised free flaps improves success rate
Authors: Smit Jeroen M. , Acosta Rafael , Zeebregts Clark J. , Liss Anders G. , Anniko Matti , Hartman ED H. M.
Published: September 2007

Introduction and aim: To develop a protocolized monitor schedule in microvascular free flap reconstruction, we investigated a possible correlation between the outcome and the interval between clamp release and start of revision. Materials and methods: All the charts of patients treated between 2000 and 2006 with a free flap were evaluated. The patients who underwent a flap revision were further analyzed. Results: A total of 608 free flaps were evaluated; 69 of these flaps were revised. Most vascular complications took place within the first 24 h; the latest complication was observed 8 days after surgery. After 6 days post surgery, the number of revisions decreased considerably. With regard to the salvaged flaps the mean time to start the revision was 46.5 h (SD 39). With regard to the failed revisions, the mean time to start the revision was 82.0 h (SD 47). This difference proved significant (P = 0.006). Conclusion: Our data shows that the majority of anastomotic failures occur within the first 24 h. Thereafter, the frequency of failures decreases. We also found that the time between initial reconstruction and start of the salvage procedure influences the outcome of the revision negatively. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
A modified model of hindlimb osteomyocutaneous flap for the study of tolerance to composite tissue allografts
Authors: Adamson Larry A. , Huang Wei‐Chao , Breidenbach Warren C. , Rahhal Dina , Xu Hong , Huang Yiming , Pidwell Diane J. , Wei Fu‐Chan , Tobin Gordon , Ildstad Suzanne T.
Published: September 2007

Composite tissue allotransplantation (CTA) is the new frontier in transplantation. More than 25 hand allograft transplants have been performed worldwide, and the feasibility has been well established. The classical experimental model of CTA involves rat orthotopic hindlimb transplantation, a time‐consuming procedure associated with high mortality and morbidity. We describe a rat heterotopic osteomyocutaneous flap that serves as a nonfunctional CTA, allowing the study of tolerance induction to a highly antigenic vascularized allograft of bone, muscle, and skin while minimizing the morbidity and mortality of full hind limb transplantation. In the present studies, we explored whether establishing chimerism by nonmyeloablative conditioning would induce tolerance to CTA. When compared with the classic hind limb transplantation model, these results demonstrate that our heterotopic hind limb flap is less morbid and as an effective experimental model for the study of CTA tolerance. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
Microsurgical arterovenous loops and biological templates: A novel in vivo chamber for tissue engineering
Authors: Manasseri Benedetto , Cuccia Giuseppe , Moimas Silvia , D'Alcontres Francesco Stagno , Polito Francesca , Bitto Alessandra , Altavilla Domenica , Squadrito Francesco , Geuna Stefano , Pattarini Lucia , Zentilin Lorena , Collesi Chiara , Puligadda Uday , Giacca Mauro , Colonna Michele Rosario
Published: September 2007

Background: Microsurgical tissue engineering is an emerging topic in regenerative medicine. Here we describe a new microsurgical model of bioengineering in rats based on the use of an arterovenous loop (AV) implanted into a commercially available crosslinked collagen/glycosaminoglycan template. Methods: The microvascular loop was created between the femoral artery and vein and covered by the template folded onto itself. The chamber was isolated from the outside tissue by an outer silicon layer to impede tissue ingrowth. Results: At 1‐month postimplantation, the tissue chamber was found heavily vascularized, as assessed by laser Doppler perfusion analysis. Histological examination showed that the AV loop was integrated into the collagen matrix of the template and that the whole template was filled with a newly formed soft connective tissue. Most interestingly, the whole scaffold was found heavily vascularized, including the formation of a large number of α‐SMA‐positive arterioles. Conclusions: The developed microsurgical chamber provides a highly vascular, isolated tool for in vivo tissue engineering. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
Free SCIA/SIEA skin flap: A dual blood supply approach to groin region
Authors: Aydin Mustafa Asim , Nasir Serdar
Published: September 2007

Free groin flap donor skin is unequaled in generous skin supply and inconspicuousness. We reliably utilize this region by taking advantage of the dual blood supply from superficial circumlex iliac (SCIA) and superficial inferior epigastric (SIEA) arteries and name the flap “Free SCIA/SIEA skin flap.” The arterial pedicle is selected between SCIA and SIEA according to vascular anatomy which is explored through an incision along the inguinal ligament prior to skin island planning. Among 57 free SCIA/SIEA skin flap transfers, two flap failures occurred; circulatory impairment was restored by taking the patient back to the operating room in four cases; major size discrepancy was noted in one end‐to‐end arterial anastomosis; and no vein grafts were required. Mostly an extended length of skin island is harvested irrespective of wound size in order to utilize the proximal skin as a vascular carrier that compensates for short pedicle. We conclude that, with the current microsurgical expertise, free SCIA/SIEA skin flap is versatile even when the skin quality is considered. © 2007 Wiley‐Liss, Inc. Microsurgery, 2007.
Separating true positive predicted residue contacts from false positive ones in mainly α proteins, using constrained Metropolis MC simulations
Authors: Vicatos Spyridon , Kaznessis Yiannis N.
Published: September 2007

We present a method that significantly improves the accuracy of predicted proximal residue pairs in protein molecules. Computational methods for predicting pairs of amino acids that are distant in the protein sequence but close in the protein 3D structure can benefit attempts to in silico recognize the fold of a protein molecule. Unfortunately, currently available methods suffer from low predictive accuracy. In this work, we use Monte Carlo simulations to fold protein molecules with proximal pair predictions used as additional energy constraints. To test our methods, we study molecules with known tertiary structures. With Monte Carlo, we generate ensembles of structures for each set of residues constraints. The distribution of the root mean square deviation of the folded structures from the known native structure reveals clear information about the accuracy of the constraint sets used. With recursive substitutions of constraints, false positive predictions are identified and filtered out and significant improvements in accuracy are observed. Proteins 2008. © 2007 Wiley‐Liss, Inc.
Extranodal Spreading of Esophageal Squamous Cell Carcinoma: Clinicopathological Characteristics and Prognostic Impact
Authors: Tanabe Tadashi , Kanda Tatsuo , Kosugi Shin‐ichi , Ikeda Yoshiyuki , Makino Shigeto , Komukai Shintaro , Ohashi Manabu , Suzuki Tsutomu , Hatakeyama Katsuyoshi
Published: September 2007

Background

Microscopic cancer spreading to extranodal connective tissues (extranodal spreading: ENS) is occasionally found in resected specimens from patients with esophageal squamous cell carcinoma (SCC), but the prognostic impact of ENS remains unclear. The aims of this study were to elucidate the prognostic impact of ENS to help determine the most suitable management for the patients with ENS.

Methods

We histologically re‐evaluated 7,349 lymph nodes obtained from 171 patients with SCC of the thoracic esophagus who underwent potentially curable resection between 1992 and 2003. We defined ENS as microscopic penetration of tumor cells from metastatic lymph nodes or tumor cell dissemination into extranodal connective tissues.

Results

Extranodal spreading was found in 37 (21.6%) patients, and it had a significant relationship with diameter and depth of the tumor, lymphatic and venous invasion, intramural metastasis, and number of metastatic nodes. Patients who were ENS positive were at higher risk of recurrence, and their overall survival rate was lower than that for ENS‐negative patients. Furthermore, recurrent risk was higher and overall survival rate was lower in ENS‐positive patients than in ENS‐negative patients when they had 1–3 metastatic nodes, but recurrent risk and overall survival rate of the patients with 4 or more metastatic nodes were very similar in ENS‐positive and ENS‐negative patients.

Conclusions

The present findings suggest that in SCC of the thoracic esophagus, the presence of ENS increases recurrent risk and reduces the overall survival of the patients with 1–3 metastatic nodes. Patients showing ENS should be managed in the same way as patients with 4 or more metastatic nodes.
The effects of maternal inflammation on neuronal development: possible mechanisms
Authors: Jonakait G. Miller
Published: September 2007

That maternal inflammation adversely affects fetal brain development is well established. Less well understood are the mechanisms that account for neurodevelopmental disorders arising from maternal inflammation. This review seeks to begin an examination of possible sites and mechanisms of action whereby inflammatory cytokines – produced by the mother or by the fetal brain – could impact the developing fetus. We focus first on the placenta where cytokines maintain the immunological environment that prevents maternal rejection of the fetus. Following a brief examination of placental transfer of maternal cytokines, the focus turns on embryonic microglia, early and ubiquitous residents of the developing brain. Finally, a more intense examination of interleukin‐6 (IL‐6) and bone morphogenetic proteins (BMPs) provides examples of glial‐ or maternal‐derived cytokines that are known to have profound effects on developing systems and that could, if dysregulated, have undesirable consequences for brain development.
Application of magnetic resonance imaging in animal models of perinatal hypoxic‐ischemic cerebral injury
Authors: Lodygensky Gregory A. , Inder Terrie E. , Neil Jeffrey J.
Published: September 2007

Brain injury occurring in the perinatal period is an important etiology of subsequent neurodevelopmental disabilities. Magnetic resonance imaging (MRI) is a tool that is used to evaluate the nature of brain injury in the human infant. MRI techniques have also been applied to various animal models of perinatal injury. The most commonly used model is the immature rat, but there have also been imaging studies in mice, rabbit kits and piglets. The studies have been carried out using MR systems of various magnetic field strengths, ranging from 1.5 to 11.7 tesla (T), with applications for quantification of infarct volume, T1 measurements, T2 measurements, proton and phosphorus spectroscopy and diffusion imaging. The MR findings are then related to histopathology and, in a few cases, behavioral evaluations. There is also a growing number of studies utilizing MRI in evaluating the efficacy of neuroprotective treatments, such as hypothermia.
Postnatal changes in 5HT and NK1 receptors in rat trigeminal motor nucleus and surroundings
Authors: Tanaka‐Gomi Nami , Yasuda Kouichi , Nakamura Masaaki , Hasumi‐Nakayama Yoko , Umemura Tetsuhiro , Tanaka Susumu , Furusawa Kiyofumi
Published: September 2007

Our previous study showed that developmental changes to serotonin and substance P coexist in the trigeminal motor nucleus (Vmo), dorsolateral subnucleus (Vmo.dl), ventromedial subnucleus (Vmo.vm) and the area within 300 μm surrounding Vmo (SVmo). This occurred in rats from embryonic day (E)19 to postnatal day (P)70, with density of these terminals peaking at P7 in these three areas. The present study examined postnatal development of serotonergic 1A receptor (5HT1A receptor) and substance P receptor (NK1 receptor) expression in Vmo and SVmo in rats from E19 to P70. No significant changes in percentages of 5HT1A and NK1 receptor‐expressing cells were seen between E19 and P70 by immunohistochemical study or in situ hybridization. In a real‐time PCR study, quantities of 5HT1A and NK1 receptor expression peaked at P7, indicating that expression of these receptors was maximized in each neuronal cell body in the Vmo and SVmo at P7. This result corresponds with postnatal changes in serotonin/substance P‐coexisting terminals found in our previous study. Furthermore, 5HT1A and NK1 receptors displayed very similar patterns of expression, which may support the hypothesis that potentiation of serotonin and substance P are involved in excitability regulating trigeminal motor functions, including mastication and breathing.
Coupling Microdialysis Sampling to Microchip Electrophoresis in a Reversibly Sealed Device
Authors: Mecker Laura C. , Martin R. Scott
Published: September 2007

In this article, we describe the fabrication and characterization of a reversibly sealed microchip device that is used to couple microdialysis sampling to microchip electrophoresis. The ability to interface microdialysis sampling and microchip electrophoresis in a device that is amenable to reversible sealing is advantageous from a repeated use standpoint. Commercially, available tubing coming from the microdialysis probe is directly inserted into the chip and flow from the probe is interfaced to the electrophoresis portion of the device through integrated pneumatic valves. Fluorescence detection was used to characterize the poly(dimethylsiloxane)-based device in terms of injection reproducibility. It was found that the entire system (microdialysis probe and microchip device) has a concentration response lag time of 170 s. Microdialysis sampling followed by an electrophoretic separation of amino acids derivatized with naphthalene-2,3-dicarboxaldehyde/cyanide was also demonstrated.
Use of a Novel Fluidics Microbead Trap/Flow-Cell Enhances Speed and Sensitivity of Bead-Based Bioassays
Authors: Ozanich Richard M. , Antolick Kate C. , Bruckner-Lea Cindy J. , Dockendorff Brian P. , Easterday Ashton N. , Edberg Heather C. , Grate Jay W. , Iyer Sujata , Johnson Laura H. , Straub Timothy M. , Valdez Cathy O. , Warner Marvin G.
Published: September 2007

Automated devices and methods for biological sample preparation often use surface functionalized microbeads (superparamagnetic or nonmagnetic) to allow capture, purification, and preconcentration of trace amounts of proteins, cells, or nucleic acids (DNA/RNA) from complex samples. We have developed unique methods and hardware for trapping either magnetic or nonmagnetic functionalized beads that allow samples and reagents to be efficiently perfused over a microcolumn of beads. This approach yields enhanced mass transport and up to fivefold improvements in assay sensitivity or speed, dramatically improving assay capability relative to assays conducted in more traditional “batch modes” (i.e., in tubes or microplate wells). Summary results are given that highlight the analytical performance improvements obtained for automated microbead processing systems using novel microbead trap/flow-cells for various applications including (1) simultaneous capture of multiple cytokines using an antibody-coupled polystyrene bead assay with subsequent flow cytometry detection; (2) capture of nucleic acids using oligonucleotide-coupled polystyrene beads with flow cytometry detection; and (3) capture of Escherichia coli 0157:H7 from 50-mL sample volumes using antibody-coupled superparamagnetic microbeads with subsequent culturing to assess capture efficiency.
Determining Dilution Accuracy in Microtiter Plate Assays Using a Quantitative Dual-Wavelength Absorbance Method
Authors: Bradshaw John Thomas , Curtis Richard H. , Knaide Tanya R. , Spaulding Benjamin W.
Published: September 2007

Dilution-based volume transfer steps are commonly performed for many types of microtiter plate-based assays. Because the exact concentrations of target samples in solution are typically not directly measured but calculated from the dilution ratio, it is important to accurately measure each dilution step so that the ultimate concentrations of those samples can be known. This paper describes a new approach, which is based on dual-dye, dual-wavelength photometry, for accurately measuring each dilution step in a multistep dilution protocol. The theory behind this method is described, as well as an experimental demonstration of the prescribed approach.
Automated Combinatorial Process for Nanofabrication of Structures Using Bioderivatized Nanoparticles
Authors: Dehlinger Dietrich , Sullivan Benjamin , Esener Sadik , Hodko Dalibor , Swanson Paul , Heller Michael J.
Published: September 2007

A fully automated electronic microarray control system (Nanochip 400 System) was used to carry out a combinatorial process to determine optimal conditions for fabricating higher order three-dimensional nanoparticle structures. Structures with up to 40 layers of bioderivatized nanoparticles were fabricated on a 400-test site CMOS microarray using the automated Nanochip 400 System. Reconfigurable electric fields produced on the surface of the CMOS microarray device actively transport, concentrate, and promote binding of 40 nm biotin- and streptavidin-derivatized nanoparticles to selected test sites on the microarray surface. The overall fabrication process including nanoparticle reagent delivery to the microarray device, electronic control of the CMOS microarray and the optical/fluorescent detection, and monitoring of nanoparticle layering are entirely controlled by the Nanochip 400 System. The automated nanoparticle layering process takes about 2 minutes per layer, with 10–20 seconds required for the electronic addressing and binding of nanoparticles, and roughly 60 seconds for washing. The addressing and building process is monitored by changes in fluorescence intensity as each nanoparticle layer is deposited. The final multilayered 3D structures are about 2 μm in thickness and 55 μm in diameter. Multilayer nanoparticle structures and control sites on the microarray were verified by SEM analysis.
A Novel Applicator Design for Contact-Free Delivery of Aqueous Reagent to Individual Biopsies in a Tissue Microarray
Authors: Gunn Dylan , Slobodan Jared , Kamran Cheyenne , Tzou Justin , Broemeling David , Banerjee Diponkar , Marziali Andre
Published: September 2007

The use of molecular techniques to inform diagnosis, prognosis, and treatment design will play an important role in the future of medicine. Each new test, however, represents a new cost to the health care system, and significant effort is required to move new techniques to the clinical setting in the most cost-effective and efficient manner. Consequently, there is a compelling need for technological improvements that will facilitate clinical application of novel assays, by reducing cost and complexity in implementation. This is particularly important in cancer pathology. Here, we present a novel applicator technology that enables staining of individual biopsies in a tissue microarray (TMA) to provide low-cost, multiplexed biomarker testing at the level of intact tissue. The applicator is designed to deliver tens of nanoliters of aqueous reagent to arrayed tissue biopsies kept under a layer of oil-based Liquid Coverslip without contacting the biopsies. A pin consisting of concentric stainless steel electrodes separated by a hydrophobic insulator provides a balance between hydrophobicity and hydrophilicity to hold a reagent droplet on the tip of the pin, whereas a small electrical current passed through the droplet spanning the electrodes is used for drop sensing. This design is more amenable to repeatable manufacturing than a previous prototype, which in initial testing demonstrated successful immunohistochemical and in situ hybridization staining of individual biopsies in a TMA, but was difficult to produce. This new design was tested to investigate the factors affecting its operation, in terms of the volume of reagent picked up and its ability to successfully deliver reagent to the biopsies.
Electrokinetic Formation of “Microbridges” for Protein Biomarkers as Sensors
Authors: Kunduru Vindhya , Prasad Shalini
Published: September 2007

We demonstrate a technique to detect protein biomarkers contained in vulnerable coronary plaque using a platform-based microelectrode array (MEA). The detection scheme is based on the property of high specificity binding between antibody and antigen similar to most immunoassay techniques. Rapid clinical diagnosis can be achieved by detecting the amount of protein in blood by analyzing the protein's electrical signature. Polystyrene beads which act as transportation agents for the immobile proteins (antigen) are electrically aligned by application of homogenous electric fields. The principle of electrophoresis is used to produce calculated electrokinetic movement among the anti-C-reactive protein (CRP), or in other words antibody funtionalized polystyrene beads. The electrophoretic movement of antibody-functionalized polystyrene beads results in the formation of “Microbridges” between the two electrodes of interest which aid in the amplification of the antigen—antibody binding event. Sensitive electrical equipment is used for capturing the amplified signal from the “Microbridge” which essentially behaves as a conducting path between the two electrodes. The technique circumvents the disadvantages of conventional protein detection methods by being rapid, noninvasive, label-free, repeatable, and inexpensive. The same principle of detection can be applied for any receptor—ligand-based system because the technique is based only on the volume of the analyte of interest. Detection of the inflammatory coronary disease biomarker CRP is achieved at concentration levels spanning over the lower microgram/milliliter to higher order nanogram/milliliter ranges.
297 Distinct Cell Lines: A High-Content Analysis Assay and a Full-Automation Design Solely Using Noncontact Liquid Dispensing
Authors: Rodriguez Richard A. , Alfonso Jaiver , O'Day Christine , Ovechkina Yulia , Ward Courtney
Published: September 2007

This review assesses the quality of the data acquired over a 13-week period from a High-Content Analysis screening project that used 297 unique cell lines. This article also evaluates the proficiency of a “tipless” (i.e., does not use disposable tips) full-automation design used for this project that prioritizes intralab system mobility and system configuration mutability. The request to assay a large number of cell lines with poorly characterized growth rates led us to devise an MDS PharmaServices, Inc. proprietary algorithm in an effort to select the proper cell plating density for each cell line. The performance metrics include coefficients of variation (CVs) of Controls for the cell plating data and Data Set Mean CVs for assessing replicate propinquity (i.e., how close the replicates are to each other). The performance of the automation system and our algorithm for this project produced data of superior quality.
Automated Technology for Performing Flow-Chemistry at Elevated Temperature and Pressure
Authors: Kovacs Ildiko , Jones Richard V. , Niesz Krisztian , Csajagi Csaba , Borcsek Bernadett , Darvas Ferenc , Urge Laszlo
Published: September 2007

Over the past few years, a series of novel microfluidic-based instruments were developed by ThalesNano, Inc. to carry out dangerous and difficult to perform chemical reactions in a safe and fast manner, resulting in superior performance to what commercial batch reactors could provide. Importance of microfluidic devices is continuously raising, as seen there are more and more publications, applications and devices in this field expanding the borders of chemistry. Furthermore, as one of the main advantages for pharmaceutical applications, these new revolutionary reactors allow the fast, on-the-fly mode optimization of different heterogeneous reactions in a high-throughput fashion. The heart of the reactor systems is the actual reactor bed, called the CatCart system. CatCarts allow easy handling of heterogeneous catalyst or immobilized reagents without further purification of products. In addition, the shoe-box size of these reactors makes them available from laboratories to industrial applications.
Microfabrication of Bilayer Polymer Actuator Valves for Controlled Drug Delivery
Authors: Tsai Han-Kuan A. , Madou Marc
Published: September 2007

Drug delivery is still a challenging mission in therapeutic treatment. Research on biomedical micro-electromechanical systems (BioMEMS) has led to a diverse range of microsystems for curative applications. This paper introduces miniaturized controlled valves and drug reservoirs for drug delivery systems. Detailed microfabrication processes, optimized package, and optical/electrochemical detection of the proposed device are described. The release mechanism of the device is controlled by a bilayer actuator valve, which consists of a conductive polymer polypyrrole (PPy) film and a thin metal gold (Au) layer. The PPy layer is electrochemically polymerized on the Au layer. Therefore, further miniaturization of the device is possible through microfabrication of the Au layer. A polydimethylsiloxane (PDMS) package is also introduced to prevent the flap from being blocked by the surrounding tissue of the human body. In addition, a parylene coating is applied to minimize the permeability of PDMS. The release process is then verified by an optical and electrochemical detection system.
Technical Considerations and Functional Outcome of 22 Major Replantations (The BSSH Douglas Lamb Lecture, 2005)
Authors: SABAPATHY S. RAJA , VENKATRAMANI H. , BHARATHI R. R. , DHEENADHAYALAN J. , BHAT V. R. , RAJASEKARAN S.
Published: September 2007

Twenty-two consecutive major replantations carried out over a 5-year period were assessed with a minimum follow-up of 2 years. Only two patients suffered guillotine amputations. The remainders were either crush, or crush avulsion amputation. Replantation was successful in 20 cases. When analysed by Chen’s criteria, there were three Grade I, nine Grade II, six Grade III and two Grade IV results. Most patients with successful replants put the hand to greater use with time and replantation greatly added to the overall well-being of the patient. We consider major replantation as a worthwhile procedure. Radical debridement, bone shortening and well laid out protocols to reduce the ischaemia time are important for success. The technical details which we believe to be important for success are outlined. With decreasing numbers of such injuries in most countries, this paper may help surgeons faced with an occasional patient with a major amputation to make the right decisions.
Uptake and spreading of anthropogenic trace gases in an eddy‐permitting model of the Atlantic Ocean
Authors: Biastoch Arne , Völker Christoph , Böning Claus W.
Published: September 2007

An eddy‐permitting circulation model of the Atlantic Ocean was used to study the effect of mesoscale processes on the uptake and spreading of anthropogenic CO2 and CFC‐11. A comparison with a coarser‐resolution model version shows anthropogenic tracer distributions with qualitatively similar patterns, but much more structure (e.g., stronger longitudinal gradients) in the eddy‐permitting model, improving the agreement with observations. The better representation of the formation of water masses such as subpolar‐mode water in the eddy‐permitting model has an influence on the distribution of anthropogenic CO2 over density classes, but no influence on the total inventory taken up. In the subpolar Atlantic, the air‐sea flux of CFC‐11 is dominated by deep‐water formation, while the air‐sea flux of anthropogenic CO2 extends over a larger part of the subpolar gyre and has a clear association with North Atlantic surface currents. An in‐depth analysis of the mechanisms shaping this distribution showed that the entrainment of water from below into the mixed layer determines the structure in the subpolar North Atlantic, whereas the temporal correlation between surface heat fluxes and mixed‐layer depth is more important in the subtropical gyre. The northward, integrated heat and anthropogenic CO2 transports in midlatitudes are closely correlated on seasonal to interannual timescales. This has implications for using the ongoing monitoring arrays of the thermohaline circulation for estimation of the transport of anthropogenic CO2.