No description available.

Neuromyelitis optica (NMO) and NMO spectrum disorders (NMOSD) are associated with the disease-specific autoantibody aquaporin-4 (AQP4-Ab), which is thought to be pathogenic. Some NMO/NMOSD patients do not have this antibody and may have different clinical and immuno-pathological disease characteristics, but previous clinical NMO/NMOSD studies have been confounded by inclusion of such patients. To define better the characteristics of AQP4-Ab disease, disease course, outcomes and predictors of disability were investigated in 106 AQP4-Ab positive NMO/NMOSD patients from the UK and Japan. AQP4-Ab positivity conferred a high risk of relapsing disease and substantial disability; age at disease onset and ethnicity were important predictors of disability type. Visual disability was more common in younger patients and those of Afro-Caribbean ethnicity whilst older patients and Caucasian patients were more at risk of motor disability. To determine whether disease characteristics were influenced by AQP4-Ab binding specificities, the differential binding of patient AQP4-Ab against the two main AQP4 isoforms was investigated. Although the relative binding to the two isoforms differed between patients, there was no association between these differences and clinical features such as relapse type, severity, onset age and ethnicity. The clinical and in vitro characteristics of AQP4-Ab negative NMO/NMOSD patients were studied. It was shown that these patients represent an aetiologically heterogeneous group. Some have other inflammatory and infectious disorders, some have low levels of AQP4-Ab and a significant proportion have antibodies to myelin oligodendrocyte glycoprotein (MOG-Ab). Others have antibodies that bind to neurons or oligodendrocytes in primary cultures. Attempts to identify novel antigens by immunocapture techniques were made, but were not successful. Patients with MOG-Ab showed differences when compared to those with AQP4-Ab including higher proportion of males, younger age at disease onset and greater likelihood of conus involvement on imaging. Additionally, patients with MOG-Ab appear to have more favourable outcomes, with better improvement from the onset attack and a lower probability of subsequent relapses. In conclusion, the work in this thesis has shown that AQP4-Ab disease is not synonymous with the term NMO and that seronegative NMO/NMOSD patients represent a clinically and aetiologically heterogeneous group and should therefore be classed separately from those with AQP4-Ab.

No description available.

Doctor of Philosophy(PhD) / The vast majority of children undergoing surgical repair of cardiac lesions do spectacularly well. However a significant proportion, ~ 25%, struggle to progress in the early postoperative period and require additional pharmacological and occasionally mechanical circulatory support. All children typically have some degree of postoperative myocardial dysfunction, with the severe spectrum termed the low cardiac output state (LCOS). LCOS is clinically defined as the requirement for new or escalated inotrope therapy, a widened arteriovenous oxygen difference, cardiac arrest or the need for reinstitution of mechanical circulatory support. LCOS is largely responsible for the morbidity and mortality involved in paediatric cardiac surgery. Despite the predictability of LCOS in the initial postoperative hours, the underlying pathophysiology remains unclear. The period of decline in cardiac function that typifies LCOS is temporally associated with the development of oedema in the tissues of the body, including the heart. This relationship between oedema and dysfunction has increasingly become blurred, with a tendency to elevate the temporal association to a causal link. We sought to explore the causes and contributions to myocardial dysfunction in this setting, including the roles of oedema and ischaemia within the heart. In focusing on oedema and ischaemia we also examined the effects of these insults on relevant myocardial membrane proteins, including those that permit rapid water transport – aquaporins (AQPs), and those involved in membrane mechanics – dystrophin, and membrane repair – dysferlin. Experimental settings which enabled the in vitro dissection of these insults and proteins of interest were combined with a clinically accurate in vivo model. This thesis describes a series of thematically linked experiments that examined LCOS, myocardial oedema and the role of various membrane proteins. We performed isolated cardiomyocyte studies, isolated heart studies as well as a clinically relevant large animal (lamb) cardiopulmonary bypass (CPB) model. Across these models we also explored the role of therapeutically protecting myocardial membranes with Poloxamer 188 (P188) and assessed any influence on myocardial function, oedema and membrane proteins. vi The results from these three models suggest that the clinically accepted dogma of a causative link between myocardial oedema and dysfunction overstates the contribution of myocardial oedema to LCOS. We found that ischaemia/reperfusion was of primary importance in causing myocardial dysfunction. Myocardial oedema without ischaemia had a mild and reversible contribution to myocardial dysfunction, but this was minor in comparison to the gross dysfunction attributable to ischaemia. Isolated cardiomyocytes, with induced oedema, functioned well. Whilst ischaemic cardiomyocytes, with less swelling still had severe contractile dysfunction. Isolated hearts, perfused with an oedema inducing crystalloid perfusate developed myocardial oedema and had minimal reversible systolic and diastolic dysfunction. Isolated hearts which experienced global ischaemia had comparable degrees of myocardial oedema, and significantly greater degrees of myocardial dysfunction that increased in severity with increasing duration of ischaemia. In the lamb CPB model, only those lambs which underwent aortic cross clamping and had a period of ischaemia had poor myocardial function. These lambs also had swollen hearts, raised myocardial AQP1 mRNA and reduced membrane dysferlin protein expression. Membrane dystrophin protein expression was not altered, somewhat unexpectedly with CPB with or without ischaemia. Lambs placed on CPB without ischaemia had good myocardial function, minimal oedema and unchanged membrane protein expression during the survival period. In a blinded lamb CPB trial of P188 there were improved haemodynamics and indicies of myocardial function associated with its use. This was also associated with preservation of dysferlin expression and reduced membrane injury. In parallel isolated heart trials of this therapy, there was a reduction in myocardial oedema associated with its use in non-ischaemic experiments. There was also a suggestion of improved diastolic function in ischaemic experiments, but no change in myocardial water content. In conclusion, we have highlighted the primacy of ischaemia/reperfusion over oedema in contributing to LCOS. We have refuted the accepted dogma that myocardial oedema causes significant dysfunction in itself, with important oedema likely to result from ischaemia. We have shown that AQP1 may be involved in the pathogenesis of the capillary leak syndrome. Finally we have hinted at a role for prophylactic P188 in the vii setting of LCOS, possibly highlighting the role of membrane repair in recovery after surgery. Isolated heart trials of P188 further support a non-rheological mechanism of action and also lend support to the causal separation of myocardial oedema and dysfunction. The integral membrane protein dysferlin, rather than dystrophin, is relevant in the setting of LCOS in the current era.

Aquaporins (AQP) are membrane bound proteins that facilitate the movement of water and other small neutral solutes across cellular membranes. Plant aquaporins belong to a large family of highly conserved proteins called the Membrane Intrinsic Protein (MIP) superfamily. In many plant species the expression of aquaporin genes and their regulation has been linked to water stress. Grapevines respond to water stress with a variety of physiological mechanisms, including the susceptibility to xylem embolism. The formation of embolised vessels can lead to a reduction in hydraulic conductivity of the xylem. Recently, it has been hypothesised that aquaporins may contribute to the water movement required for embolism recovery of xylem vessels thus restoring the hydraulic pathway. Molecular and physiological techniques have been combined to study the putative role of plasma membrane and tonoplast membrane aquaporins in response to water stress induced xylem embolism in two cultivars of grapevine (Vitis vinifera cv. Chardonnay and Grenache). Water-stress induced cavitation was measured in the stems and petioles of pot grown grapevines of a drought tolerant (Grenache) and a drought sensitive variety (Chardonnay) by the detection of ultrasonic acoustic emissions (UAEs) over both a drying and diurnal cycle. Vulnerability curves were generated by correlating the UAEs with the leaf water potential (ψL). Varietal differences in cavitation vulnerability and hydraulic properties were observed. Grenache was more susceptible to water-stress induced xylem embolism than Chardonnay, and displayed a higher hydraulic capacity (measured by maximum hydraulic conductivity). This is most likely due to anatomical differences of the xylem vessels. Chardonnay displayed vulnerability segmentation, with cavitation occurring first in the petiole and later in the stem, before developing into “runaway” cavitation under severe water stress. Vulnerability segmentation was not observed in Grenache, with both petioles and stems equally vulnerable to the formation of xylem embolism. Under severe water stress, Grenache did not develop runaway cavitation indicating that they must have some mechanism to prevent the onset of runaway cavitation. To determine the role of aquaporins, candidate genes were identified, by screening a Vitis vinifera cv. Cabernet Sauvignon cDNA library, for aquaporin cDNAs encoding members of the Plasma membrane Intrinsic Protein (PIP) and Tonoplast Intrinsic Protein (TIP) subfamilies. The screen resulted in the identification of 11 full-length and two partial aquaporin cDNAs. Sequence analyses of these cDNAs reveal five are homologous to PIP2 aquaporins, six to PIP1 and two to the TIP aquaporins. Functional expression of the fulllength AQP cDNAs in Xenopus oocytes showed PIP2 members have significantly higher water permeability compared to PIP1 aquaporins. VvPIP2;1 showed very high water permeability which was reduced by acidic cytosolic pH, as has been reported for other members of the PIP2 family. Transcript analysis of some of these aquaporin genes provides preliminary evidence that aquaporins may contribute to differences in the hydraulic response of these two grapevine varieties to conditions of water stress. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1313316 / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2008

Doctor of Philosophy(PhD) / The vast majority of children undergoing surgical repair of cardiac lesions do spectacularly well. However a significant proportion, ~ 25%, struggle to progress in the early postoperative period and require additional pharmacological and occasionally mechanical circulatory support. All children typically have some degree of postoperative myocardial dysfunction, with the severe spectrum termed the low cardiac output state (LCOS). LCOS is clinically defined as the requirement for new or escalated inotrope therapy, a widened arteriovenous oxygen difference, cardiac arrest or the need for reinstitution of mechanical circulatory support. LCOS is largely responsible for the morbidity and mortality involved in paediatric cardiac surgery. Despite the predictability of LCOS in the initial postoperative hours, the underlying pathophysiology remains unclear. The period of decline in cardiac function that typifies LCOS is temporally associated with the development of oedema in the tissues of the body, including the heart. This relationship between oedema and dysfunction has increasingly become blurred, with a tendency to elevate the temporal association to a causal link. We sought to explore the causes and contributions to myocardial dysfunction in this setting, including the roles of oedema and ischaemia within the heart. In focusing on oedema and ischaemia we also examined the effects of these insults on relevant myocardial membrane proteins, including those that permit rapid water transport – aquaporins (AQPs), and those involved in membrane mechanics – dystrophin, and membrane repair – dysferlin. Experimental settings which enabled the in vitro dissection of these insults and proteins of interest were combined with a clinically accurate in vivo model. This thesis describes a series of thematically linked experiments that examined LCOS, myocardial oedema and the role of various membrane proteins. We performed isolated cardiomyocyte studies, isolated heart studies as well as a clinically relevant large animal (lamb) cardiopulmonary bypass (CPB) model. Across these models we also explored the role of therapeutically protecting myocardial membranes with Poloxamer 188 (P188) and assessed any influence on myocardial function, oedema and membrane proteins. vi The results from these three models suggest that the clinically accepted dogma of a causative link between myocardial oedema and dysfunction overstates the contribution of myocardial oedema to LCOS. We found that ischaemia/reperfusion was of primary importance in causing myocardial dysfunction. Myocardial oedema without ischaemia had a mild and reversible contribution to myocardial dysfunction, but this was minor in comparison to the gross dysfunction attributable to ischaemia. Isolated cardiomyocytes, with induced oedema, functioned well. Whilst ischaemic cardiomyocytes, with less swelling still had severe contractile dysfunction. Isolated hearts, perfused with an oedema inducing crystalloid perfusate developed myocardial oedema and had minimal reversible systolic and diastolic dysfunction. Isolated hearts which experienced global ischaemia had comparable degrees of myocardial oedema, and significantly greater degrees of myocardial dysfunction that increased in severity with increasing duration of ischaemia. In the lamb CPB model, only those lambs which underwent aortic cross clamping and had a period of ischaemia had poor myocardial function. These lambs also had swollen hearts, raised myocardial AQP1 mRNA and reduced membrane dysferlin protein expression. Membrane dystrophin protein expression was not altered, somewhat unexpectedly with CPB with or without ischaemia. Lambs placed on CPB without ischaemia had good myocardial function, minimal oedema and unchanged membrane protein expression during the survival period. In a blinded lamb CPB trial of P188 there were improved haemodynamics and indicies of myocardial function associated with its use. This was also associated with preservation of dysferlin expression and reduced membrane injury. In parallel isolated heart trials of this therapy, there was a reduction in myocardial oedema associated with its use in non-ischaemic experiments. There was also a suggestion of improved diastolic function in ischaemic experiments, but no change in myocardial water content. In conclusion, we have highlighted the primacy of ischaemia/reperfusion over oedema in contributing to LCOS. We have refuted the accepted dogma that myocardial oedema causes significant dysfunction in itself, with important oedema likely to result from ischaemia. We have shown that AQP1 may be involved in the pathogenesis of the capillary leak syndrome. Finally we have hinted at a role for prophylactic P188 in the vii setting of LCOS, possibly highlighting the role of membrane repair in recovery after surgery. Isolated heart trials of P188 further support a non-rheological mechanism of action and also lend support to the causal separation of myocardial oedema and dysfunction. The integral membrane protein dysferlin, rather than dystrophin, is relevant in the setting of LCOS in the current era.

BACKGROUND: The accumulation of misfolded proteins is a hallmark of many neurodegenerative disorders, including Chronic Traumatic Encephalopathy (CTE). Intracellular protein degradation pathways appear to be insufficient in preventing or halting disease progression. A brain-wide waste clearance pathway mediated by astroglial aquaporin-4 (AQP4) water channels in the perivascular space called the “glymphatic system” has recently been identified. Disruption of this system due to mislocalization of AQP4 away from perivascular astrocytic endfeet (“depolarization”) is linked to reductions in solute clearance and the build up of toxic metabolites in different neurologic conditions associated with aging and traumatic brain injury. Accumulation of aggregated tau protein around blood vessels at the depths of cortical sulci is considered the pathognomonic lesion of CTE, and may reflect impairment of glymphatic pathway function in these perivascular spaces. OBJECTIVES: To investigate whether changes in AQP4 expression or perivascular AQP4 polarization are present in CTE and to assess their relationship with CTE lesions. Additionally, AQP4 expression in CTE will be compared to subjects with a pathological diagnosis of Alzheimer’s disease (AD) and non-pathological controls without a history of head trauma. METHODS: Postmortem frontal cortex samples from neuropatholigcally confirmed cases of CTE, AD, and non-pathological controls were provided by the VA-BU-CLF Brain Bank. Fixed tissue samples were cut at 20 microns from each case and immunofluorescently stained for AQP4, glial fibrillary acidic protein (GFAP), and phosphorylated tau (AT-8). Slides were imaged using a Zeiss 880 Airyscan confocal microscope and analyzed using the HALO image software analysis platform. RESULTS: Increased perivascular AQP4 polarization was significantly associated with lesional vessels compared to non-lesional vessels in CTE (p=0.0187). When assessed between groups, CTE showed less AQP4 polarization surrounding non-lesional vessels compared to controls, and seemingly higher polarization around lesional vessels compared to AD, however these differences were not statistically significant. CONCLUSIONS: Blood brain barrier (BBB) breakdown is a common occurrence following traumatic brain injury (TBI) and has previously been confirmed in postmortem cases of CTE. The findings reported in the current study showing increased, rather than decreased, perivascular AQP4 polarization around lesional vessels compared to non-lesional vessels in CTE may therefore reflect a compensatory mechanism of astrocytes in response to secondary vasogenic edema in the face of chronic inflammation and disrupted BBB integrity, rather than acute cytotoxic edema which is likely the main driver of AQP4 depolarization reported in previous studies.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第20255号 / 医博第4214号 / 新制||医||1020(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 渡邉 大, 教授 萩原 正敏, 教授 影山 龍一郎 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2006. / Härtill 4 uppsatser.

Hintergrund: Die bakterielle Meningitis hat eine Letalität von 10-20%. Das Hirnödem stellt bei ca. 14 % der Erkrankten eine prognosebestimmende Komplikation dar. Ein aktueller Forschungsansatz umfasst die Bedeutung der Aquaporine für die Entwicklung, Aufrechterhaltung und Resorption der verschiedenen Hirnödemformen, insbesondere des zytotoxischen und des vasogenen Hirnödems. In dieser Arbeit wird untersucht, ob Aquaporin 1 und Aquaporin 4 im Liquor von Patienten mit bakterieller und viraler Meningitis, im Vergleich zu einer gesunden Kontrollgruppe, nachweisbar sind. Zudem sollte geklärt werden, ob sich hieraus eine differenzialdiagnostische Einordnung ergibt und sich Rückschlüsse auf das Ausmaß eines Hirnödems und das Outcome schließen lassen. Methode: Aquaporin 1 und 4 wurde im Liquor und im Serum von Patienten mit bakterieller (nCSF = 35 , nSerum = 20) und viraler  (nCSF = 22) Meningitis sowie in einer Kontrollgruppe (nCSF = 27 , nSerum = 12) mittels eines (kommerziell erhältlichen) ELISAs bestimmt. Klinische Daten und Routinelaborparameter wurden verglichen und in Korrelation zu den Aquaporinkonzentrationen gesetzt. Ergänzend wurde bei einer Untergruppe der Patienten mit bakterieller Meningitis (n = 8) eine neuropsychologische Testung durchgeführt. Ergebnisse: Aquaporin 1 und 4 ließen sich in allen Gruppen nachweisen, ca. 40% der Aquaporin 4 Konzentrationen lagen unterhalb der Nachweisgrenze des ELISAs. Im Gruppenvergleich aller drei Gruppen unterschieden sich die Aquaporin 1-Konzentrationen (p = 0,0001) und die Aquaporin 4-Konzentrationen (p = 0,035) im Liquor signifikant voneinander. In der Gruppe der Patienten mit bakterieller Meningitis ließ sich eine negative Korrelation zwischen Aquaporin 1 und 4 im Liquor feststellen (r = - 0,519, p = 0,002). Aussagekräftige Korrelationen der klinischen Daten, der liquor- und laborchemischen Parameter sowie der neuropsychologischen Testergebnisse zu den Aquaporin 1- und Aquaporin 4-Konzentrationen fanden sich nicht.  Diskussion: In dieser Arbeit konnte erstmalig gezeigt werden, dass Aquaporin 1 und Aquaporin 4 im Liquor (und Serum) von Patienten mit einer bakteriellen und viralen Meningitis sowie in einer Kontrollgruppe nachweisbar sind. Für Aquaporin 1 und Aquaporin 4 im Liquor fanden sich signifikante Unterschiede im Vergleich aller Gruppen im Kruskal-Wallis-Test. Rückschlüsse bezüglich einer differenzial-diagnostischen Einordnung zur viralen Meningitis konnten nicht gezogen werden. Aussagen zur Schwere eines Hirnödems und zur Prognose können mit den vorliegenden Daten nicht getroffen werden. Der Ursprung der gemessenen Aquaporine bei Patienten mit Meningitis lässt sich in dieser Arbeit nicht abschließend klären und bedarf weiterer Grundlagenforschung.