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1.  Scapholunate Ligament Reconstruction 
Journal of Wrist Surgery  2013;2(2):110-115.
Background Scapholunate reconstruction poses a challenge to orthopedic surgeons.
Materials and Methods Prospective cohort.
Description of Technique Our technique for scapholunate (SL) reconstruction involves ligament reconstruction utilizing a portion of the flexor carpi radialis tendon rerouted via transosseous tunnels across the scaphoid, lunate, and triquetrum (scapholunotriquetral tenodesis). The tendon graft is secured with interference screw fixation into the triquetrum. The philosophy of this new technique is to reduce subluxation and maintain the relationship between scaphoid and lunate by placing a graft through the center of the SL articulation. This graft is then tensioned by passing it centrally through the lunate and triquetrum and secured using an interference screw in the triquetrum. Secondary stabilizers, including the dorsal intercarpal ligament, are then augmented by passing the graft back to the scaphoid, crossing from the triquetrum over the proximal capitate. This further reinforces the translational relationship between the scaphoid and the triquetrum and, therefore, augments stability of the SL articulation.
Results We have utilized this technique successfully in over 40 patients since 2009. We report on a prospective consecutive series of 11 patients with over 12 months follow-up (range 12 to 24 months) demonstrating good early radiological and clinical outcomes.
Conclusions In developing this technique, we aimed to take the best features of previously described techniques and address the perceived shortcomings of each. We believe there are several benefits of our technique. Moreover, few other techniques address as many of the aspects of chronic SL instability as our technique does.
doi:10.1055/s-0033-1341962
PMCID: PMC3699271  PMID: 24436802
scapholunate; luno-triquetral; reconstruction; wrist; interference screw; surgical technique
2.  Stainless steel versus titanium volar multi-axial locking plates for fixation of distal radius fractures: a randomised clinical trial 
Background
Distal radius fractures are among the most common fractures seen in the hospital emergency department. Of these, over 40% are considered unstable and require some form of fixation. In recent years with the advent of low profile plating, open reduction and internal fixation (ORIF) using volar plates has become the surgical treatment of choice in many hospitals. However, it is currently unknown which plating system has the lowest complication rate and/or superior clinical and radiological outcomes following surgery. Few studies have compared different types of plates, which may have various features, different plate and screw designs or may be manufactured from different materials (for example, stainless steel or titanium). This study will specifically investigate and compare the clinical and radiological outcomes and complication rates of two commonly used volar plating systems for fixation of distal radius fractures: one made from stainless steel (Trimed™ Volar Plate, Trimed™, California, USA) and the other made from titanium (Medartis® Aptus Volar Plate, Medartis®, Basel, Switzerland). The primary aim of this study is to determine if there is a difference on the Patient Reported Wrist Evaluation six months following ORIF using a volar plate for adult patients with a distal radius fracture.
Methods/Design
This study will implement a randomized prospective clinical trial study design evaluating the outcomes of two different types of volar plates: one plate manufactured from stainless steel (Trimed™ Volar Plate) and one plate manufactured from titanium (Medartis® Aptus Volar Plate). The surgery will be performed at a major trauma hospital in Brisbane, Australia. Outcome measures including function, adverse events, range of movement, strength, disability, radiological findings and health-related quality of life will be collected at 6 weeks, 3, 6, 12 and 24 months following surgery. A parallel economic analysis will also be performed. This randomized clinical trial is due to deliver results in December 2016.
Discussion
Results from this trial will contribute to the evidence on operative management of distal radius fractures and plate material type.
Trial registration
ACTRN12612000969864
doi:10.1186/1471-2474-15-74
PMCID: PMC3984716  PMID: 24612524
Distal radius; Randomized trial; Volar plating; Trimed; Medartis; Wrist; Surgery
3.  Cellular, pharmacological, and biophysical evaluation of explanted lungs from a patient with sickle cell disease and severe pulmonary arterial hypertension 
Pulmonary Circulation  2013;3(4):936-951.
Pulmonary hypertension is recognized as a leading cause of morbidity and mortality in patients with sickle cell disease (SCD). We now report benchtop phenotyping from the explanted lungs of the first successful lung transplant in SCD. Pulmonary artery smooth muscle cells (PASMCs) cultured from the explanted lungs were analyzed for proliferate capacity, superoxide (O2•−) production, and changes in key pulmonary arterial hypertension (PAH)–associated molecules and compared with non-PAH PASMCs. Upregulation of several pathologic processes persisted in culture in SCD lung PASMCs in spite of cell passage. SCD lung PASMCs showed growth factor– and serum-independent proliferation, upregulation of matrix genes, and increased O2•− production compared with control cells. Histologic analysis of SCD-associated PAH arteries demonstrated increased and ectopically located extracellular matrix deposition and degradation of elastin fibers. Biomechanical analysis of these vessels confirmed increased arterial stiffening and loss of elasticity. Functional analysis of distal fifth-order pulmonary arteries from these lungs demonstrated increased vasoconstriction to an α1-adrenergic receptor agonist and concurrent loss of both endothelial-dependent and endothelial-independent vasodilation compared with normal pulmonary arteries. This is the first study to evaluate the molecular, cellular, functional, and mechanical changes in end-stage SCD-associated PAH.
doi:10.1086/674754
PMCID: PMC4070844  PMID: 25006410
sickle cell disease; pulmonary arterial hypertension; lung transplant; CD47; thrombospondin 1; endothelin 1; superoxide; matrix
4.  BAP1 loss defines a new class of renal cell carcinoma 
Nature genetics  2012;44(7):751-759.
The molecular pathogenesis of renal cell carcinoma (RCC) is poorly understood. Whole-genome and exome sequencing followed by innovative tumorgraft analyses (to accurately determine mutant allele ratios) identified several putative two-hit tumor suppressor genes including BAP1. BAP1, a nuclear deubiquitinase, is inactivated in 15% of clear-cell RCCs. BAP1 cofractionates with and binds to HCF-1 in tumorgrafts. Mutations disrupting the HCF-1 binding motif impair BAP1-mediated suppression of cell proliferation, but not H2AK119ub1 deubiquitination. BAP1 loss sensitizes RCC cells in vitro to genotoxic stress. Interestingly, BAP1 and PBRM1 mutations anticorrelate in tumors (P=3×10−5), and combined loss of BAP1 and PBRM1 in a few RCCs was associated with rhabdoid features (q=0.0007). BAP1 and PBRM1 regulate seemingly different gene expression programs, and BAP1 loss was associated with high tumor grade (q=0.0005). Our results establish the foundation for an integrated pathological and molecular genetic classification of RCC, paving the way for subtype-specific treatments exploiting genetic vulnerabilities.
doi:10.1038/ng.2323
PMCID: PMC3788680  PMID: 22683710
5.  Activated VEGF Receptor Shed Into the Vitreous in Eyes With Wet AMD 
Archives of ophthalmology  2009;127(5):613-621.
Objective
To examine whether phosphorylated vascular endothelial growth factor (VEGF) receptors shed into the vitreous reflect the ongoing retinal and choroidal signal pathway activity in wet age-related macular degeneration (AMD).
Methods
Vitreous samples obtained immediately prior to anti-VEGF injection from 11 patients with choroidal neovascularization were analyzed using reverse-phase microarrays. Two patients had samples collected at the time of injection and 1 month later. Samples from 5 patients were collected prior to vitrectomy for macular hole, epiretinal membrane, or retinal detachment.
Results
Phosphorylated forms of VEGF receptor (VEGFR Y996 and Y1175), platelet-derived growth factor receptor β (PDGFRβ Y716 and Y751), and c-KIT (Y703) were present in the vitreous. A significant difference in PDGFRβ Y751 (P <.002), VEGFR Y996 (P <.04), and VEGFR Y1175 (P <.006), but not c-KIT Y703 (P <.05) or PDGFRβ Y716 (P <.96), was noted for the responders to treatment (n=5) compared with nonresponders (n=6) and controls (n=5).
Conclusions
Vitreous levels of activated receptors constitute a new class of biomarkers. Activated forms of VEGF and PDGF receptors, previously not known to exist in the vitreous, correlate with response to anti-VEGF therapy. These findings could provide the basis for the development of individualized treatment and discovery of new therapeutic targets.
doi:10.1001/archophthalmol.2009.88
PMCID: PMC3777775  PMID: 19433709
6.  A Proteomic Analysis of Eccrine Sweat: Implications for the Discovery of Schizophrenia Biomarker Proteins 
Journal of proteome research  2012;11(4):2127-2139.
Liquid chromatography tandem mass spectrometry (LC-MS/MS) and multiple reaction monitoring mass spectrometry (MRM-MS) proteomics analyses were performed on eccrine sweat of healthy controls, and the results were compared with those from individuals diagnosed with schizophrenia (SZ). This is the first large scale study of the sweat proteome. First, we performed LC-MS/MS on pooled SZ samples and pooled control samples for global proteomics analysis. Results revealed a high abundance of diverse proteins and peptides in eccrine sweat. Most of the proteins identified from sweat samples were found to be different than the most abundant proteins from serum, which indicates that eccrine sweat is not simply a plasma transudate, and may thereby be a source of unique disease-associated biomolecules. A second independent set of patient and control sweat samples were analyzed by LC-MS/MS and spectral counting to determine qualitative protein differential abundances between the control and disease groups. Differential abundances of selected proteins, initially determined by spectral counting, were verified by MRM-MS analyses. Seventeen proteins showed a differential abundance of approximately two-fold or greater between the SZ pooled sample and the control pooled sample. This study demonstrates the utility of LC-MS/MS and MRM-MS as a viable strategy for the discovery and verification of potential sweat protein disease biomarkers.
doi:10.1021/pr2007957
PMCID: PMC3703649  PMID: 22256890
Sweat; Proteome; Schizophrenia; Biomarkers; LC-MS/MS; MRM
7.  The Heme Degradation Pathway is a Promising Serum Biomarker Source for the Early Detection of Alzheimer's Disease 
Journal of Alzheimer's disease : JAD  2010;19(3):1081-1091.
One of the remaining challenges in Alzheimer's disease (AD) research is the establishment of biomarkers for early disease detection. As part of a prospective study spanning a period of five years, we have collected serial serum samples from cognitively normal, mild cognitively impaired (MCI), and mild AD participants, including same patient samples before and after cognitive decline. Using mass spectrometry we identified several promising leads for biomarker development, such as prosaposin, phospholipase D1, biliverdin reductase B, and S100 calcium binding protein A7. Selected candidate markers were verified using reverse phase protein microarray assays. Of 15 protein/protein abundance ratios that were significantly altered in sera from subjects with mild AD compared to Normal or MCI subjects, 14 were composed of ratios containing heme oxygenase-1, biliverdin reductase A, or biliverdin reductase B. Moreover, an increase in the protein abundance ratio of matrix metallopeptidase 9/biliverdin reductase differentiated stable MCI subjects from MCI subjects progressing into mild AD before the onset of cognitive decline. These findings strongly implicate the heme degradation pathway as a promising source of protein biomarkers for the early detection of AD.
doi:10.3233/JAD-2010-1303
PMCID: PMC3589529  PMID: 20157261
Alzheimer's disease; biomarker; BLVR; BVR; complement factor H; heme; heme oxyengase-1; phospholipase D1; prosaposin; S100A7; serum
8.  A Validated Tumorgraft Model Reveals Activity of Dovitinib Against Renal Cell Carcinoma 
Science translational medicine  2012;4(137):137ra75.
Most anticancer drugs entering clinical trials fail to achieve approval from the US FDA. Drug development is hampered by the lack of preclinical models with therapeutic predictive value. Herein, we report the development and validation of a tumorgraft model of renal cell carcinoma (RCC) and its application to the evaluation of an experimental drug. Tumor samples from 94 patients were implanted in the kidney of mice without additives or disaggregation. Tumors from 35 patients formed tumorgrafts, and 16 stable lines were established. Samples from metastatic sites engrafted at high frequency, and stable engraftment of primary tumors in mice correlated with decreased patient survival suggesting that tumor growth in mice may reveal the acquisition by the tumor of an ability to thrive at distant sites and metastasize. Tumorgrafts retained the histology, gene expression, DNA copy number alterations, and over 90% of the protein-coding gene mutations of the corresponding tumors. As determined by the induction of hypercalcemia in tumorgraft-bearing mice, tumorgrafts were able to act on the host causing paraneoplastic syndromes. In studies simulating drug exposures in patients, RCC tumorgraft growth was inhibited by sunitinib and sirolimus (into which temsirolimus is converted in humans), but not by erlotinib, which was used as a control. Dovitinib, a drug in clinical development, showed greater activity than sunitinib and sirolimus. The routine incorporation of models recapitulating the molecular genetics and drug sensitivities of human tumors into preclinical programs has the potential to improve oncology drug development.
doi:10.1126/scitranslmed.3003643
PMCID: PMC3570965  PMID: 22674553
xenograft; tumor graft; orthotopic; PK; NOD/SCID; kidney cancer; clear-cell renal cell carcinoma
9.  A mouse model of accelerated liver aging due to a defect in DNA repair 
Hepatology (Baltimore, Md.)  2012;55(2):609-621.
The liver changes with age leading to an impaired ability to respond to hepatic insults and increased incidence of liver disease in the elderly. Therefore, there is critical need for rapid model systems to study aging-related liver changes. One potential opportunity is murine models of human progerias, or diseases of accelerated aging. Ercc1−/Δ mice model a rare human progeroid syndrome caused by inherited defects in DNA repair. To determine if hepatic changes that occur with normal aging occur prematurely in Ercc1−/Δ mice, we systematically compared liver from 5 month-old, progeroid Ercc1−/Δ mice to old (24–36 month) wild-type (WT) mice. Both displayed areas of necrosis, foci of hepatocellular degeneration and acute inflammation. Loss of hepatic architecture, fibrosis, steatosis, pseudocapillarization, and anisokaryosis were more dramatic in Ercc1−/Δ mice than in old WT mice. Liver enzymes were significantly elevated in serum of Ercc1−/Δ mice and old WT mice, while albumin was reduced, demonstrating liver damage and dysfunction. The regenerative capacity of Ercc1−/Δ liver following partial hepatectomy was significantly reduced. There was evidence of increased oxidative damage in Ercc1−/Δ and old WT liver, including lipofuscin, lipid hydroperoxides and acrolein as well as increased hepatocellular senescence. There was a highly significant correlation in genome-wide transcriptional changes between old WT and 16 but not 5 week-old Ercc1−/Δ mice emphasizing that the Ercc1−/Δ mice acquire an aging profile in early adulthood.
Conclusion
There are strong functional, regulatory and histopathological parallels between accelerated aging driven by a DNA repair defect and normal aging. This supports a role for DNA damage in driving aging and validates a murine model for rapidly testing hypotheses about causes and treatment for aging-related hepatic changes.
doi:10.1002/hep.24713
PMCID: PMC3250572  PMID: 21953681
DNA repair; DNA damage; progeria; hepatic fibrosis; mouse model
10.  Effects of Carpal Tunnel Syndrome on Dexterous Manipulation Are Grip Type-Dependent 
PLoS ONE  2013;8(1):e53751.
Carpal tunnel syndrome (CTS) impairs sensation of a subset of digits. Although the effects of CTS on manipulation performed with CTS-affected digits have been studied using precision grip tasks, the extent to which CTS affects multi-digit force coordination has only recently been studied. Whole-hand manipulation studies have shown that CTS patients retain the ability to modulate multi-digit forces to object mass, mass distribution, and texture. However, CTS results in sensorimotor deficits relative to healthy controls, including significantly larger grip force and lower ability to balance the torques generated by the digits. Here we investigated the effects of CTS on multi-digit force modulation to object weight when manipulating an object with a variable number of fingers. We hypothesized that CTS patients would be able to modulate digit forces to object weight. However, as different grip types involve the exclusive use of CTS-affected digits (‘uniform’ grips) or a combination of CTS-affected and non-affected digits (‘mixed’ grips), we addressed the question of whether ‘mixed’ grips would reduce or worsen CTS-induced force coordination deficits. The former scenario would be due to adding digits with intact tactile feedback, whereas the latter scenario might occur due to a potentially greater challenge for the central nervous system of integrating ‘noisy’ and intact tactile feedback. CTS patients learned multi-digit force modulation to object weight regardless of grip type. Although controls exerted the same total grip force across all grip types, patients exerted significantly larger grip force than controls but only for manipulations with four and five digits. Importantly, this effect was due to CTS patients’ inability to change the finger force distribution when adding the ring and little fingers. These findings suggest that CTS primarily challenges sensorimotor integration processes for dexterous manipulation underlying the coordination of CTS-affected and non-affected digits.
doi:10.1371/journal.pone.0053751
PMCID: PMC3542366  PMID: 23326498
11.  Accurate Whole Human Genome Sequencing using Reversible Terminator Chemistry 
Bentley, David R. | Balasubramanian, Shankar | Swerdlow, Harold P. | Smith, Geoffrey P. | Milton, John | Brown, Clive G. | Hall, Kevin P. | Evers, Dirk J. | Barnes, Colin L. | Bignell, Helen R. | Boutell, Jonathan M. | Bryant, Jason | Carter, Richard J. | Cheetham, R. Keira | Cox, Anthony J. | Ellis, Darren J. | Flatbush, Michael R. | Gormley, Niall A. | Humphray, Sean J. | Irving, Leslie J. | Karbelashvili, Mirian S. | Kirk, Scott M. | Li, Heng | Liu, Xiaohai | Maisinger, Klaus S. | Murray, Lisa J. | Obradovic, Bojan | Ost, Tobias | Parkinson, Michael L. | Pratt, Mark R. | Rasolonjatovo, Isabelle M. J. | Reed, Mark T. | Rigatti, Roberto | Rodighiero, Chiara | Ross, Mark T. | Sabot, Andrea | Sankar, Subramanian V. | Scally, Aylwyn | Schroth, Gary P. | Smith, Mark E. | Smith, Vincent P. | Spiridou, Anastassia | Torrance, Peta E. | Tzonev, Svilen S. | Vermaas, Eric H. | Walter, Klaudia | Wu, Xiaolin | Zhang, Lu | Alam, Mohammed D. | Anastasi, Carole | Aniebo, Ify C. | Bailey, David M. D. | Bancarz, Iain R. | Banerjee, Saibal | Barbour, Selena G. | Baybayan, Primo A. | Benoit, Vincent A. | Benson, Kevin F. | Bevis, Claire | Black, Phillip J. | Boodhun, Asha | Brennan, Joe S. | Bridgham, John A. | Brown, Rob C. | Brown, Andrew A. | Buermann, Dale H. | Bundu, Abass A. | Burrows, James C. | Carter, Nigel P. | Castillo, Nestor | Catenazzi, Maria Chiara E. | Chang, Simon | Cooley, R. Neil | Crake, Natasha R. | Dada, Olubunmi O. | Diakoumakos, Konstantinos D. | Dominguez-Fernandez, Belen | Earnshaw, David J. | Egbujor, Ugonna C. | Elmore, David W. | Etchin, Sergey S. | Ewan, Mark R. | Fedurco, Milan | Fraser, Louise J. | Fajardo, Karin V. Fuentes | Furey, W. Scott | George, David | Gietzen, Kimberley J. | Goddard, Colin P. | Golda, George S. | Granieri, Philip A. | Green, David E. | Gustafson, David L. | Hansen, Nancy F. | Harnish, Kevin | Haudenschild, Christian D. | Heyer, Narinder I. | Hims, Matthew M. | Ho, Johnny T. | Horgan, Adrian M. | Hoschler, Katya | Hurwitz, Steve | Ivanov, Denis V. | Johnson, Maria Q. | James, Terena | Jones, T. A. Huw | Kang, Gyoung-Dong | Kerelska, Tzvetana H. | Kersey, Alan D. | Khrebtukova, Irina | Kindwall, Alex P. | Kingsbury, Zoya | Kokko-Gonzales, Paula I. | Kumar, Anil | Laurent, Marc A. | Lawley, Cynthia T. | Lee, Sarah E. | Lee, Xavier | Liao, Arnold K. | Loch, Jennifer A. | Lok, Mitch | Luo, Shujun | Mammen, Radhika M. | Martin, John W. | McCauley, Patrick G. | McNitt, Paul | Mehta, Parul | Moon, Keith W. | Mullens, Joe W. | Newington, Taksina | Ning, Zemin | Ng, Bee Ling | Novo, Sonia M. | O'Neill, Michael J. | Osborne, Mark A. | Osnowski, Andrew | Ostadan, Omead | Paraschos, Lambros L. | Pickering, Lea | Pike, Andrew C. | Pike, Alger C. | Pinkard, D. Chris | Pliskin, Daniel P. | Podhasky, Joe | Quijano, Victor J. | Raczy, Come | Rae, Vicki H. | Rawlings, Stephen R. | Rodriguez, Ana Chiva | Roe, Phyllida M. | Rogers, John | Rogert Bacigalupo, Maria C. | Romanov, Nikolai | Romieu, Anthony | Roth, Rithy K. | Rourke, Natalie J. | Ruediger, Silke T. | Rusman, Eli | Sanches-Kuiper, Raquel M. | Schenker, Martin R. | Seoane, Josefina M. | Shaw, Richard J. | Shiver, Mitch K. | Short, Steven W. | Sizto, Ning L. | Sluis, Johannes P. | Smith, Melanie A. | Sohna, Jean Ernest Sohna | Spence, Eric J. | Stevens, Kim | Sutton, Neil | Szajkowski, Lukasz | Tregidgo, Carolyn L. | Turcatti, Gerardo | vandeVondele, Stephanie | Verhovsky, Yuli | Virk, Selene M. | Wakelin, Suzanne | Walcott, Gregory C. | Wang, Jingwen | Worsley, Graham J. | Yan, Juying | Yau, Ling | Zuerlein, Mike | Rogers, Jane | Mullikin, James C. | Hurles, Matthew E. | McCooke, Nick J. | West, John S. | Oaks, Frank L. | Lundberg, Peter L. | Klenerman, David | Durbin, Richard | Smith, Anthony J.
Nature  2008;456(7218):53-59.
doi:10.1038/nature07517
PMCID: PMC2581791  PMID: 18987734
12.  HEPATOCYTE GAMMA-CATENIN COMPENSATES FOR CONDITIONALLY DELETED BETA-CATENIN AT ADHERENS JUNCTIONS 
Journal of hepatology  2011;55(6):1256-1262.
Background and Aims
Wnt/β-catenin signaling is important in liver physiology. β-Catenin is also pivotal in adherens junctions (AJ). Here, we investigate hepatocyte-specific β-catenin-conditional null mice (KO), for any alterations in AJ and related tight junctions (TJ).
Materials and Methods
Using genearray, PCR, western blot, immunohistochemistry, immunofluorescence and coprecipitation studies, we compare and contrast the composition of AJ and TJ in KO and littermate wild-type (WT) control livers.
Results
We demonstrate E-cadherin-β-catenin association in epithelial cells in WT livers, which is lost in the KOs. While total levels of α-catenin, E-cadherin and F-actin were modestly decreased, KO livers show increased γ-catenin/plakoglobin. By co-precipitation, E-cadherin-β-catenin-F-actin association was observed in WT livers, but E-cadherin-γ-catenin-F-actin association was evident in KO livers. γ-Catenin was localized at the hepatocyte membrane at baseline in the KO liver. While γ-catenin gene expression remained unaltered, an increase in serine- and threonine-phosphorylated, but not tyrosine-phosphorylated γ-catenin was observed in KO livers by immunoprecipitation. A continued presence of γ-catenin at hepatocyte membrane without any nuclear localization was observed in liver regeneration after partial-hepatectomy at 40 and 72 hours in both KO and WT by immunofluorescence. Analysis of TJ revealed lack of claudin-2 and increased levels of JAM-A and claudin-1 in KO livers.
Conclusions
γ-Catenin adequately maintains AJ in the absence of β-catenin in hepatocytes, however it lacks nuclear localization. Also, β-catenin-claudin-2 may be an important mechanism of crosstalk between the AJ and TJ.
doi:10.1016/j.jhep.2011.03.014
PMCID: PMC3221911  PMID: 21703193
13.  CARBON MONOXIDE INDUCES HYPOTHERMIA TOLERANCE IN KUPFFER CELLS AND ATTENUATES LIVER ISCHEMIA/REPERFUSION INJURY IN RATS 
Ischemia/reperfusion (I/R) injury in liver grafts, initiated by cold preservation and augmented by reperfusion, is a major problem complicating graft quality, post-transplant patient care, and outcomes of liver transplantation (LTx). Kupffer cells (KC) play important roles in I/R injury; however, little is known about their changes during cold preservation. We examined whether pretreatment with carbon monoxide (CO), a cytoprotective product of heme degradation, would influence KC activity during cold storage and protect the liver graft against LTx-induced I/R injury. In vitro, primary rat KC were stimulated for 24 hrs with hypothermia (4°C, 20% O2), LPS, or hypoxia (37°C, 5% O2) with and without CO pretreatment. When exposed to hypothermia, rat KC produced ROS, but not TNF-α or NO. Preincubation of KC with CO upregulated HSP70 and inhibited ROS generation. When liver grafts obtained from donor rats exposed to CO (250 ppm) for 24 hrs were transplanted after 18 hrs cold preservation in UW solution, HSP70 expression in the grafts increased, and serum AST/ALT levels as well as necrotic area and inflammatory infiltrates were significantly reduced after LTx, when compared to control grafts. CO-pretreated liver grafts showed less TNF-α, ICAM-1 and iNOS mRNA upregulation, as well as reduced pro-apoptotic Bax mRNA, cleaved caspase-3 and PARP expressions. Thus, donor pretreatment with CO ameliorates I/R injury associated with LTx, with an increased hepatic HSP70 expression, particularly in KC population.
doi:10.1002/lt.22415
PMCID: PMC3222745  PMID: 21850691
14.  Effects of Carpal Tunnel Syndrome on adaptation of multi-digit forces to object mass distribution for whole-hand manipulation 
Background
Carpal tunnel syndrome (CTS) is a compression neuropathy of the median nerve that results in sensorimotor deficits in the hand. Until recently, the effects of CTS on hand function have been studied using mostly two-digit grip tasks. The purpose of this study was to investigate the coordination of multi-digit forces as a function of object center of mass (CM) during whole-hand grasping.
Methods
Fourteen CTS patients and age- and gender-matched controls were instructed to grasp, lift, hold, and release a grip device with five digits for seven consecutive lifts while maintaining its vertical orientation. The object CM was changed by adding a mass at different locations at the base of the object. We measured forces and torques exerted by each digit and object kinematics and analyzed modulation of these variables to object CM at object lift onset and during object hold. Our task requires a modulation of digit forces at and after object lift onset to generate a compensatory moment to counteract the external moment caused by the added mass and to minimize object tilt.
Results
We found that CTS patients learned to generate a compensatory moment and minimized object roll to the same extent as controls. However, controls fully exploited the available degrees of freedom (DoF) in coordinating their multi-digit forces to generate a compensatory moment, i.e., digit normal forces, tangential forces, and the net center of pressure on the finger side of the device at object lift onset and during object hold. In contrast, patients modulated only one of these DoFs (the net center of pressure) to object CM by modulating individual normal forces at object lift onset. During object hold, however, CTS patients were able to modulate digit tangential force distribution to object CM.
Conclusions
Our findings suggest that, although CTS did not affect patients’ ability to perform our manipulation task, it interfered with the modulation of specific grasp control variables. This phenomenon might be indicative of a lower degree of flexibility of the sensorimotor system in CTS to adapt to grasp task conditions.
doi:10.1186/1743-0003-9-83
PMCID: PMC3543219  PMID: 23171737
Sensorimotor memories; Grasping; Learning; Center of mass
15.  The loss of renal dendritic cells and activation of host adaptive immunity are long-term effects of ischemia/reperfusion injury following syngenic kidney transplantation 
Kidney International  2012;81(10):1015-1025.
Ischemia/reperfusion injury associated with kidney transplantation induces profound acute injury, influences early graft function and affects long-term graft outcomes. To determine whether renal dendritic cells play any role during initial innate ischemia/reperfusion injury and the subsequent development of adaptive immune responses, we studied the behavior and function of renal graft and host infiltrating dendritic cells during early and late phases of renal ischemia/reperfusion injury. Wild type to GFP-transgenic rat kidney transplantation was performed with and without 24 hours cold storage. Ischemia/reperfusion injury in cold stored grafts resulted in histopathological changes of interstitial fibrosis and tubular atrophy by 10 weeks accompanied by upregulation of mRNAs of mediators of interstitial fibrosis and inflammation. In normal rat kidneys we identified two populations of renal dendritic cells, predominant CD103−CD11b/c+ and minor CD103+CD11b/c+ cells. After transplantation without cold storage, grafts maintained CD103− but not CD103+ GFP-negative renal dendritic cells for 10 weeks. In contrast, both cell subsets disappeared from cold stored grafts, which associated with a significant GFP-expressing host CD11b/c+ cell infiltration that included CD103+ dendritic cells with a TNF-α producing phenotype. These changes in graft/host dendritic cell populations were associated with progressive infiltration of host CD4+ T cells with effector/effector-memory phenotypes and IFN-γ secretion. Thus, renal graft ischemia/reperfusion injury causes graft dendritic cell loss and was associated with progressive host dendritic cell and T cell recruitment. Renal resident dendritic cells might function as a protective regulatory network.
doi:10.1038/ki.2011.458
PMCID: PMC3340432  PMID: 22278023
CD103; IF/TA; effector CD4+ T cells; IFN-α; TNF-α
16.  Endogenous Histones Function as Alarmins in Sterile Inflammatory Liver Injury through Toll-like Receptor 9 
Hepatology (Baltimore, Md.)  2011;54(3):999-1008.
Sterile inflammatory insults are known to activate innate immunity and propagate organ damage through the recognition of extracellular Damage Associated Molecular Pattern (DAMP) molecules. Although DAMPs, such as endogenous DNA and nuclear High Mobility Group Box 1, have been shown to be critical in sterile inflammation, the role of nuclear histone proteins has not yet been investigated. We report that endogenous histones function as DAMPs following ischemic injury through the pattern recognition receptor Toll-Like Receptor 9 (TLR9) to initiate inflammation. Using an in vivo model of hepatic ischemia/reperfusion (I/R) injury, we show that levels of circulating histones are significantly higher after I/R, and that histone neutralization significantly protects against injury. Injection of exogenous histones exacerbates I/R injury through cytotoxic effects mediated by TLR9 and MyD88. In addition, histone administration increases TLR9 activation, while neither TLR9 nor MyD88 mutant mice respond to exogenous histones. Furthermore, we demonstrate in vitro that extracellular histones enhance DNA-mediated TLR9 activation in immune cells through a direct interaction.
Conclusions
these novel findings reveal that histones represent a new class of DAMP molecules and they serve as a crucial link between initial damage and activation of innate immunity during sterile inflammation.
doi:10.1002/hep.24501
PMCID: PMC3213322  PMID: 21721026
Sterile Inflammation; Ischemia/reperfusion injury; innate immunity; danger associated molecular pattern molecules
17.  Hepatic B7-H1 expression is essential to control cold ischemia/reperfusion injury after mouse liver transplantation 
Hepatology (Baltimore, Md.)  2011;54(1):216-228.
Ischemia/reperfusion (I/R) injury remains a key risk factor significantly affecting morbidity and mortality after liver transplantation (LTx). B7-H1, recently identified member of the B7 family, is known to play important roles in regulating local immune responses. We hypothesized that B7-H1 plays crucial roles during innate immune responses induced by hepatic I/R injury and tested this hypothesis in the mouse LTx model using B7-H1 KO liver grafts with 24 hr cold storage. Cold I/R injury in WT to WT LTx enhanced constitutive B7-H1 expression on dendritic cells and sinusoidal endothelial cells, and promptly induced B7-H1 on hepatocytes. When B7-H1 KO liver grafts were transplanted into WT recipients, serum ALT levels and graft necrosis were significantly higher than WT to WT LTx. Augmented tissue injury in B7-H1 KO grafts was associated with increased frequencies and absolute numbers of graft CD3+ T cells, in particular CD8+ T cells. B7-H1 KO grafts had significantly lower incidences of Annexin V+ CD8+ T cells, indicating the failure to delete infiltrating CD8+ T cells. To evaluate the relative contribution of parenchymal and bone marrow-derived cell (BMDC) B7-H1 expression, chimeric liver grafts lacking B7-H1 on parenchymal cells or BMDC were generated and transplanted into WT recipients. Selective B7-H1 deficiency on parenchymal cells or BMDC resulted in similar levels of ALT and liver injury, suggesting that both parenchymal and BMDC B7-H1expression is involved in the control of liver damage. Human livers upregulated B7-H1 expression after LTx. Conclusion: The study demonstrates that graft tissue expression of B7-H1 plays critical roles in regulating inflammatory responses during LTx-induced hepatic I/R injury, and suggests that negative coregulatory signals may have an important function in hepatic innate immune responses.
doi:10.1002/hep.24360
PMCID: PMC3125416  PMID: 21503939
apoptosis; T cells; hepatocyte; nonparenchymal cells; dendritic cells
18.  Head injury (moderate to severe) 
Clinical Evidence  2010;2010:1210.
Introduction
Head injury in young adults is often associated with motor vehicle accidents, violence, and sports injuries. In older adults it is often associated with falls. Severe head injury can lead to secondary brain damage from cerebral ischaemia resulting from hypotension, hypercapnia, and raised intracranial pressure. Severity of brain injury is assessed using the Glasgow Coma Scale (GCS). While about one quarter of people with severe brain injury (GCS score less than 8) will make a good recovery, about one third will die, and one fifth will have severe disability or be in a vegetative state.
Methods and outcomes
We conducted a systematic review and aimed to answer the following clinical question: What are the effects of interventions to reduce complications of moderate to severe head injury as defined by Glasgow Coma Scale? We searched: Medline, Embase, The Cochrane Library, and other important databases up to November 2009 (Clinical Evidence reviews are updated periodically, please check our website for the most up-to-date version of this review). We included harms alerts from relevant organisations such as the US Food and Drug Administration (FDA) and the UK Medicines and Healthcare products Regulatory Agency (MHRA).
Results
We found 17 systematic reviews, RCTs, or observational studies that met our inclusion criteria.
Conclusions
In this systematic review we present information relating to the effectiveness and safety of the following interventions: antibiotics, anticonvulsants, corticosteroids, hyperventilation, hypothermia, and mannitol.
Key Points
Head injury in young adults is often associated with motor vehicle accidents, violence, and sports injuries. In older adults it is often associated with falls. This review covers only moderate to severe head injury. Severe head injury can lead to secondary brain damage from cerebral ischaemia resulting from hypotension, hypercapnia, and raised intracranial pressure.Poor outcome correlates with low post-resuscitation Glasgow Coma Scale (GCS) score, older age, eye pupil abnormalities, hypoxia or hypotension before definitive treatment, traumatic subarachnoid haemorrhage, and inability to control intracranial pressure.Severity of brain injury is assessed using the GCS. While about one quarter of people with severe brain injury (GCS score less than 8) will make a good recovery, about one third will die, and one fifth will have severe disability or be in a vegetative state.
There is no strong evidence of benefit from any treatment in reducing the complications of moderate to severe head injury. Despite this, most clinicians implement various combinations of treatments discussed here.
Hyperventilation and mannitol are frequently used to lower intracranial pressure. Anticonvulsants, barbiturates, antibiotics, and hypothermia are less commonly implemented. Evidence on hyperventilation, mild hypothermia, and mannitol has been inconclusive. Carbamazepine and phenytoin may reduce early seizures in people with head injury, but they have not been shown to reduce late seizures, neurological disability, or death.Barbiturates have not been shown to be effective in reducing intracranial pressure or in preventing adverse neurological outcomes after head injury.Prophylactic antibiotics have not been shown to reduce the risk of death or meningitis in people with skull fracture.
CAUTION: Corticosteroids have been shown to increase mortality when used acutely in people with head injury. One large RCT (the CRASH trial) found that death from all causes and severe disability at 6 months were more likely in people with head injury given methylprednisolone infusion than in those given placebo. Corticosteroids are no longer used in the treatment of head injuries.
PMCID: PMC3217652  PMID: 21418686
19.  A Whole-Organ Regenerative Medicine Approach for Liver Replacement 
Background & Aims
The therapy of choice for end-stage liver disease is whole-organ liver transplantation, but this option is limited by a shortage of donor organs. Cell-based therapies and hepatic tissue engineering have been considered as alternatives to liver transplantation, but neither has proven effective to date. A regenerative medicine approach for liver replacement has recently been described that includes the use of a three-dimensional organ scaffold prepared by decellularization of xenogeneic liver. The present study investigates a new, minimally disruptive method for whole-organ liver decellularization and three different cell reseeding strategies to engineer functional liver tissue.
Methods
A combination of enzymatic, detergent, and mechanical methods are used to remove all cells from isolated rat livers. Whole-organ perfusion is used in a customized organ chamber and the decellularized livers are examined by morphologic, biochemical, and immunolabeling techniques for preservation of the native matrix architecture and composition. Three different methods for hepatocyte seeding of the resultant three-dimensional liver scaffolds are evaluated to maximize cell survival and function: (1) direct parenchymal injection, (2) multistep infusion, or (3) continuous perfusion.
Results
The decellularization process preserves the three-dimensional macrostructure, the ultrastructure, the composition of the extracellular matrix components, the native microvascular network of the liver, and the bile drainage system, and up to 50% of growth factor content. The three-dimensional liver matrix reseeded with the multistep infusion of hepatocytes generated ∼90% of cell engraftment and supported liver-specific functional capacities of the engrafted cells, including albumin production, urea metabolism, and cytochrome P450 induction.
Conclusions
Whole-organ liver decellularization is possible with maintenance of structure and composition suitable to support functional hepatocytes.
doi:10.1089/ten.tec.2010.0698
PMCID: PMC3103054  PMID: 21375407
20.  Effects of Carpal Tunnel Syndrome on Adaptation of Multi-Digit Forces to Object Weight for Whole-Hand Manipulation 
PLoS ONE  2011;6(11):e27715.
The delicate tuning of digit forces to object properties can be disrupted by a number of neurological and musculoskeletal diseases. One such condition is Carpal Tunnel Syndrome (CTS), a compression neuropathy of the median nerve that causes sensory and motor deficits in a subset of digits in the hand. Whereas the effects of CTS on median nerve physiology are well understood, the extent to which it affects whole-hand manipulation remains to be addressed. CTS affects only the lateral three and a half digits, which raises the question of how the central nervous system integrates sensory feedback from affected and unaffected digits to plan and execute whole-hand object manipulation. We addressed this question by asking CTS patients and healthy controls to grasp, lift, and hold a grip device (445, 545, or 745 g) for several consecutive trials. We found that CTS patients were able to successfully adapt grip force to object weight. However, multi-digit force coordination in patients was characterized by lower discrimination of force modulation to lighter object weights, higher across-trial digit force variability, the consistent use of excessively large digit forces across consecutive trials, and a lower ability to minimize net moments on the object. Importantly, the mechanical requirement of attaining equilibrium of forces and torques caused CTS patients to exert excessive forces at both CTS-affected digits and digits with intact sensorimotor capabilities. These findings suggest that CTS-induced deficits in tactile sensitivity interfere with the formation of accurate sensorimotor memories of previous manipulations. Consequently, CTS patients use compensatory strategies to maximize grasp stability at the expense of exerting consistently larger multi-digit forces than controls. These behavioral deficits might be particularly detrimental for tasks that require fine regulation of fingertip forces for manipulating light or fragile objects.
doi:10.1371/journal.pone.0027715
PMCID: PMC3218012  PMID: 22110738
21.  A comprehensive catalogue of somatic mutations from a human cancer genome 
Nature  2009;463(7278):191-196.
All cancers carry somatic mutations. A subset of these somatic alterations, termed driver mutations, confer selective growth advantage and are implicated in cancer development, whereas the remainder are passengers. Here we have sequenced the genomes of a malignant melanoma and a lymphoblastoid cell line from the same person, providing the first comprehensive catalogue of somatic mutations from an individual cancer. The catalogue provides remarkable insights into the forces that have shaped this cancer genome. The dominant mutational signature reflects DNA damage due to ultraviolet light exposure, a known risk factor for malignant melanoma, whereas the uneven distribution of mutations across the genome, with a lower prevalence in gene footprints, indicates that DNA repair has been preferentially deployed towards transcribed regions. The results illustrate the power of a cancer genome sequence to reveal traces of the DNA damage, repair, mutation and selection processes that were operative years before the cancer became symptomatic.
doi:10.1038/nature08658
PMCID: PMC3145108  PMID: 20016485
22.  Investigation of the Ovarian and Prostate Cancer Peptidome for Candidate Early Detection Markers Using a Novel Nanoparticle Biomarker Capture Technology 
The AAPS Journal  2010;12(4):504-518.
Current efforts to identify protein biomarkers of disease use mainly mass spectrometry (MS) to analyze tissue and blood specimens. The low-molecular-weight “peptidome” is an attractive information archive because of the facile nature by which the low-molecular-weight information freely crosses the endothelial cell barrier of the vasculature, which provides opportunity to measure disease microenvironment-associated protein analytes secreted or shed into the extracellular interstitium and from there into the circulation. However, identifying useful protein biomarkers (peptidomic or not) which could be useful to detect early detection/monitoring of disease, toxicity, doping, or drug abuse has been severely hampered because even the most sophisticated, high-resolution MS technologies have lower sensitivities than those of the immunoassays technologies now routinely used in clinical practice. Identification of novel low abundance biomarkers that are indicative of early-stage events that likely exist in the sub-nanogram per milliliter concentration range of known markers, such as prostate-specific antigen, cannot be readily detected by current MS technologies. We have developed a new nanoparticle technology that can, in one step, capture, concentrate, and separate the peptidome from high-abundance blood proteins. Herein, we describe an initial pilot study whereby the peptidome content of ovarian and prostate cancer patients is investigated with this method. Differentially abundant candidate peptidome biomarkers that appear to be specific for early-stage ovarian and prostate cancer have been identified and reveal the potential utility for this new methodology
doi:10.1208/s12248-010-9211-3
PMCID: PMC2977006  PMID: 20549403
Peptidome; Cancer; Biomarker; Nanoparticle; Mass spectrometry
23.  EX VIVO APPLICATION OF CARBON MONOXIDE IN UW SOLUTION PREVENTS TRANSPLANT-INDUCED RENAL ISCHEMIA/REPERFUSION INJURY IN PIGS 
I/R injury is a major deleterious factor of successful kidney transplantation (KTx). Carbon monoxide (CO) is an endogenous gaseous regulatory molecule, and exogenously delivered CO in low concentrations provides potent cytoprotection. This study evaluated efficacies of CO exposure to excised kidney grafts to inhibit I/R injury in the pig KTx model. Porcine kidneys were stored for 48 hrs in control UW or UW supplemented with CO (CO-UW) and autotransplanted in a 14-day follow-up study. In the control UW group, animal survival was 80% (4/5) with peak serum creatinine levels of 12.0±5.1 mg/dl. CO-UW showed potent protection, and peak creatinine levels were reduced to 6.9±1.4 mg/dl with 100% (5/5) survival without any noticeable adverse event or abnormal COHb value. Control grafts at 14d showed significant tubular damages, focal fibrotic changes, and numerous infiltrates. The CO-UW group showed significantly less severe histopathological changes with less TGF-β and p-Smad3 expression. Grafts in CO-UW also showed significantly lower early mRNA levels for proinflammatory cytokines and less lipid peroxidation. CO in UW provides significant protection against renal I/R injury in the porcine KTx model. Ex vivo exposure of kidney grafts to CO during cold storage may therefore be a safe strategy to reduce I/R injury.
doi:10.1111/j.1600-6143.2010.03040.x
PMCID: PMC2886983  PMID: 20199500
24.  An Initial Characterization of the Serum Phosphoproteome 
Journal of proteome research  2009;8(12):5523-5531.
Phosphorylation is a dynamic post-translational protein modification that is the basis of a general mechanism for maintaining and regulating protein structure and function, and of course underpins key cellular processes through signal transduction. In the last several years, many studies of large-scale profiling of phosphoproteins and mapping phosphorylation sites from cultured human cells or tissues by mass spectrometry technique have been published; however, there is little information on general (or global) phosphoproteomic characterization and description of the content of phosphoprotein analytes within the circulation. Circulating phosphoproteins and phosphopeptides could represent important disease biomarkers because of their well-known importance in cellular function, and these analytes frequently are mutated and activated in human diseases such as cancer. Here we report an initial attempt to characterize the phosphoprotein content of serum. To accomplish this, we developed a method in which phosphopeptides are enriched from digested serum proteins and analyzed by LC-MS/MS using LTQ-Orbitrap (CID) and LTQ-ETD mass spectrometers. Using this approach we identified ~100 unique phosphopeptides with stringent filtering criteria and a lower than 1% false discovery rate.
doi:10.1021/pr900603n
PMCID: PMC2789176  PMID: 19824718
Titanium dioxide; phosphopeptide; serum; LTQ-Orbitrap; LTQ-ETD; mass spectrometry
25.  LIVER GRAFT EXPOSURE TO CARBON MONOXIDE DURING COLD STORAGE PROTECTS SINUSOIDAL ENDOTHELIAL CELLS AND AMELIORATES REPERFUSION INJURY IN RATS 
Background
Hepatic ischemia/reperfusion (I/R) injury significantly influences short and long-term outcomes after liver transplantation (LTx). The critical step initiating the injury is known to include sinusoidal endothelial cell (SEC) alteration during the cold preservation period. As carbon monoxide (CO) has potent cytoprotective functions on vascular endothelial cells, this study examines if CO treatment of excised liver grafts during cold storage could protect SEC and ameliorate hepatic I/R injury.
Methods
Rat liver grafts were preserved in UW solution containing 5% CO (CO-UW) for 18–24 hrs and transplanted into syngenic Lewis rats.
Results
After 18 hrs cold preservation, SEC damage was evident with Propidium Iodide (PI) nuclear stain on SEC, and the frequency of PI+ SEC was significantly lower in grafts stored in CO-UW than in control UW. SEC protection with CO associated with decreased ICAM translocation and less MMP release during cold preservation. After LTx with 18 hrs cold preservation, serum ALT levels and hepatic necrosis were significantly less in CO-UW than in control UW group. With 24 hrs cold storage, 35% (7/20) survived with control UW, while the survival with CO-UW improved to 80% (8/10). These beneficial effects of CO-UW associated with significant reduction of neutrophil extravasation, down-regulation of hepatic mRNA for TNF-α and ICAM-1, and less hepatic ERK activation. Liver grafts from Kupffer cell depleted donors or pseudogerm-free donors showed less SEC death during cold preservation, and CO-UW further reduced SEC death.
Conclusion
CO delivery to excised liver grafts during cold preservation efficiently ameliorates SEC damage and hepatic I/R injury.
doi:10.1002/lt.21918
PMCID: PMC2930486  PMID: 19877256
ex vivo treatment; ischemia/reperfusion; liver transplantation

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