Effects of a human recombinant alkaline phosphatase during impaired mitochondrial function in human renal proximal tubule epithelial cells

Sepsis-associated acute kidney injury is a multifactorial syndrome in which inflammation and renal microcirculatory dysfunction play a profound role. Subsequently, renal tubule mitochondria reprioritize cellular functions to prevent further damage. Here, we investigated the putative protective effects of human recombinant alkaline phosphatase (recAP) during inhibition of mitochondrial respiration in conditionally immortalized human proximal tubule epithelial cells (ciPTEC). Full inhibition of mitochondrial oxygen consumption was obtained after 24h antimycin A treatment, which did not affect cell viability. While recAP did not affect the antimycin A-induced decreased oxygen consumption and increased hypoxia-inducible factor-1α or adrenomedullin gene expression levels, the antimycin A-induced increase of pro-inflammatory cytokines IL-6 and IL-8 was attenuated. Antimycin A tended to induce the release of detrimental purines ATP and ADP, which reached statistical significance when antimycin A was co-incubated with lipopolysaccharide, and were completely converted into cytoprotective adenosine by recAP. As the adenosine A2A receptor was up-regulated after antimycin A exposure, an adenosine A2A receptor knockout ciPTEC cell line was generated in which recAP still provided protection. Together, recAP did not affect oxygen consumption but attenuated the inflammatory response during impaired mitochondrial function, an effect suggested to be mediated by dephosphorylating ATP and ADP into adenosine

An Experimental Study to Measure Grout Penetrability, Improve the Grout Spread, and Evaluate the Real Time Grouting Control Theory

Due to the significant influence of the grout penetrability properties on spread of grout in rock fractures, this study aimed to investigate the grout penetrability from four different aspects. In Part (a), after review of all the existing methodologies developed to measure the grout penetrability, Filter-pump and Penetrability-meter were examined against Short-slot to figure out which one is more reliable. The study decisively considered Short-slot more reliable. In part (b), the so-called varying aperture long slot (VALS), an artificial fracture with apertures of 230-10 μm, was developed to study the gout penetrability more realistically. In part (c), a low-frequency rectangular pressure impulse was introduced to improve the grout spread by successive erosion of the produced filter cakes in consecutive cycles. The results showed considerable improvement in experiments using Short-slot. The dissipation of the pressure impulses was then investigated using VALS with noticeable remaining amplitudes after 2.0-2.7 m. In part (d), VALS was once more introduced to examine RTGC theory in a fracture with variable aperture. The study showed a relatively satisfactory agreement between the experimental results and the predictions of the grout propagation using the hydraulic aperture, whereas the predictions using the mean physical aperture showed considerably faster spread.För att uppnå den täthet som krävs i undermarkskonstruktioner är det nödvändigt att uppnå tillräcklig spridning av bruket vid injekteringen. Cementbaserade injekteringsmedel är vanligast inom injekteringsindustrin, eftersom det har flera fördelar, särskilt från ekonomisk och miljömässig synvinkel. Eftersom inträngningsförmågan kan påverkas betydligt hos injekteringsmedel beroende på vilket bruk man använder, är syftet med denna avhandling att studera: a) Vilka av de befintliga metoder som har utvecklats för att mäta inträngningsförmågan hos injekteringsmedel är tillförlitliga? b) Hur kan man mäta inträngningsförmågan hos injekteringsmedel mer realistiskt? c) Hur kan man förbättra spridningen hos injekteringsmedel med hjälp av dynamiska tryckimpulser? och d) Kan Real Time Grouting Control (RTGC)-teorin användas för att förutse spridningen hos injekteringsmedel i en artificiell spricka med varierande vidd? I del a) av studien genomfördes en undersökning av alla befintliga metoder som har utvecklats för att mäta inträngningsförmågan hos injekteringsmedel. Därefter genomfördes en jämförelse mellan Filterpumpen, Filterpressen (d.v.s. två av de vanligaste metoderna i svensk injekteringsindustri) och metoden Kort spalt under så lika provförhållanden som möjligt. Studien visade att Kort spalt är tillförlitligare på grund av dess mer realistiska provförhållanden (d.v.s. geometrin, trycket och injekteringsvolymen) och är därmed en bättre utvärderingsmetod. I del b) utvecklades en så kallad Varying Aperture Long Slot (VALS), en fyra meter lång artificiell spricka med minskande spaltvidder (från 230 till 10 μm), som är en mer realistisk metod för att studera inträngningsförmågan hos injekteringsmedel under statiska/dynamiska tryckförhållanden upp till 20 bar. I del c) användes en lågfrekvent rektangulär tryckimpuls för att förbättra spridningen hos injekteringsmedlet genom successiv erosion av filterkakor som har byggts vid förträngningar i konsekutiva cykler. Resultaten visade en förbättring på upp till 11 gånger mer volym i mätningar med Kort spalt med 30-43 μm breda spaltvidder. Sedan, spridningen av tryckimpulserna undersöktes längs VALS. Resultaten visade att de återstående amplituderna av tryckimpulser kan vara så stora som 46% respektive 25% av den applicerade amplituden 2,0 m respektive 2,7 m in i sprickan. I del d) användes VALS igen för att undersöka om RTGC-teorin kan användas för att bedöma spridningen av injekteringsmedel i en konstgjord spricka med variabel spaltvidd. Studien visade en förhållandevis tillfredsställande överensstämmelse mellan försöksresultaten och förutsägelserna av spridningen hos injekteringsmedel när man tog hänsyn till den hydrauliska öppningen. Som jämföreslse gav förutsägelserna baserade på den genomsnittliga fysiska öppningen (felaktigt) en betydligt snabbare spridning. Detta visar att användning av den genomsnittliga fysiska öppningen inte alltid är lämpligt vid tillämpning av RTGC-teori. Beroende på de geometriska förhållandena kan den hydrauliska öppningen ge en mer realistisk förutsägelse av spridningen hos injekteringsmedlet.QC 20171121</p

Human scattered tubular cells represent a heterogeneous population of glycolytic dedifferentiated proximal tubule cells

Scattered tubular cells (STCs) are a phenotypically distinct cell population in the proximal tubule that increase in number after acute kidney injury. We aimed to characterize the human STC population. Three-dimensional human tissue analysis revealed that STCs are preferentially located within inner bends of the tubule and are barely present in young kidney tissue (<2 years), and their number increases with age. Increased STC numbers were associated with acute tubular injury (kidney injury molecule 1) and interstitial fibrosis (alpha smooth muscle actin). Isolated CD13+CD24−CD133− proximal tubule epithelial cells (PTECs) and CD13+CD24+ and CD13+CD133+ STCs were analyzed using RNA sequencing. Transcriptome analysis revealed an upregulation of nuclear factor κB, tumor necrosis factor alpha, and inflammatory pathways in STCs, whereas metabolism, especially the tricarboxylic acid cycle and oxidative phosphorylation, was downregulated, without showing signs of cellular senescence. Using immunostaining and a publicly available single-cell sequencing database of human kidneys, we demonstrate that STCs represent a heterogeneous population in a transient state. In conclusion, STCs are dedifferentiated PTECs showing a metabolic shift toward glycolysis, which could facilitate cellular survival after kidney injury

Human scattered tubular cells represent a heterogeneous population of glycolytic dedifferentiated proximal tubule cells

Scattered tubular cells (STCs) are a phenotypically distinct cell population in the proximal tubule that increase in number after acute kidney injury. We aimed to characterize the human STC population. Three-dimensional human tissue analysis revealed that STCs are preferentially located within inner bends of the tubule and are barely present in young kidney tissue (<2 years), and their number increases with age. Increased STC numbers were associated with acute tubular injury (kidney injury molecule 1) and interstitial fibrosis (alpha smooth muscle actin). Isolated CD13+CD24−CD133− proximal tubule epithelial cells (PTECs) and CD13+CD24+ and CD13+CD133+ STCs were analyzed using RNA sequencing. Transcriptome analysis revealed an upregulation of nuclear factor κB, tumor necrosis factor alpha, and inflammatory pathways in STCs, whereas metabolism, especially the tricarboxylic acid cycle and oxidative phosphorylation, was downregulated, without showing signs of cellular senescence. Using immunostaining and a publicly available single-cell sequencing database of human kidneys, we demonstrate that STCs represent a heterogeneous population in a transient state. In conclusion, STCs are dedifferentiated PTECs showing a metabolic shift toward glycolysis, which could facilitate cellular survival after kidney injury