Depletion of ATP in renal tubular cells plays the central role in the pathogenesis of kidney diseases. Nevertheless, inability to visualize spatiotemporal
ATP distribution and dynamics has hindered ...further analysis.
A novel mouse line systemically expressing an ATP biosensor (an ATP synthase subunit and two fluorophores) revealed spatiotemporal ATP dynamics at single-cell resolution during warm and cold ischemic reperfusion (IR) with two-photon microscopy. This experimental system enabled quantification of fibrosis 2 weeks after IR and assessment of the relationship between the ATP recovery in acute phase and fibrosis in chronic phase.
Upon ischemia induction, the ATP levels of proximal tubule (PT) cells decreased to the nadir within a few minutes, whereas those of distal tubule (DT) cells decreased gradually up to 1 hour. Upon reperfusion, the recovery rate of ATP in PTs was slower with longer ischemia. In stark contrast, ATP in DTs was quickly rebounded irrespective of ischemia duration. Morphologic changes of mitochondria in the acute phase support the observation of different ATP dynamics in the two segments. Furthermore, slow and incomplete ATP recovery of PTs in the acute phase inversely correlated with fibrosis in the chronic phase. Ischemia under conditions of hypothermia resulted in more rapid and complete ATP recovery with less fibrosis, providing a proof of concept for use of hypothermia to protect kidney tissues.
Visualizing spatiotemporal ATP dynamics during IR injury revealed higher sensitivity of PT cells to ischemia compared with DT cells in terms of energy metabolism. The ATP dynamics of PTs in AKI might provide prognostic information.
A World Health Organization (WHO) guideline-based multimodal hand hygiene (HH) initiative was introduced hospital-wide to a nonteaching Japanese hospital for 5 years. The objective of this study was ...to assess the effect of this initiative in terms of changes in alcohol-based hand rub (ABHR) consumption and the Hand Hygiene Self-Assessment Framework (HHSAF) score.
The consumption of monthly hospital-wide ABHR was calculated in L per 1000 patient days (PDs). The change in ABHR consumption was analysed by an interrupted time series analysis with a pre-implementation period of 36 months and an implementation period of 60 months. The correlation between annual ABHR consumption and the HHSAF score was estimated using Pearson's correlation coefficients.
The annual ABHR consumption was 4.0 (L/1000 PDs) to 4.4 in the pre-implementation period and 10.4 to 34.4 in the implementation period. The HHSAF score was 117.5 (out of 500) in the pre-implementation period and 267.5 to 445 in the implementation period. A statistically significant increase in the monthly ABHR consumption (change in slope: + 0.479 L/1000 PDs, p < 0.01) was observed with the implementation of the initiative. Annual ABHR consumption was strongly correlated with the annual HHSAF score (r = 0.971, p < 0.01).
A 5-year WHO-based HH initiative significantly increased ABHR consumption. Our study suggested that the HHSAF assessment can be a good process measure to improve HH in a single facility, as ABHR consumption increased with the HHSAF score.
The role of hepatic ATP-binding cassette transporter 1 (ABCA1) in maintaining plasma high density lipoprotein cholesterol (HDL-C) levels is well established, but its role in reverse cholesterol ...transport (RCT) is unclear. Probucol is a compound that reduces HDL-C levels but also reduces atherosclerosis in animal models and xanthomas in humans. The aim of the present study was to test the hypothesis that probucol inhibits hepatic ABCA1 activity, thereby reducing HDL-C levels but promoting RCT from macrophages.
Wild-type (WT) C57BL/6 mice and scavenger receptor class B type I (SR-BI) knockout mice were fed a chow diet containing 0.5% probucol or normal chow for 2 weeks. In WT mice, probucol, despite decreasing HDL-C by >80%, effectively maintained macrophage RCT. In SR-BI knockout mice, probucol also substantially reduced HDL-C but significantly increased macrophage RCT. Furthermore, probucol significantly enhanced the excretion of HDL-derived cholesterol into feces in both WT and SR-BI knockout mice. Probucol inhibited ABCA1-dependent cholesterol efflux from mouse primary hepatocytes, and this effect was shown to be responsible for the effect of probucol on increasing the fecal excretion of HDL-derived cholesterol in vivo.
We demonstrate that pharmacological inhibition of hepatic ABCA1 activity with probucol reduced HDL-C levels but promoted RCT through diversion of HDL-derived cholesterol from efflux back into plasma instead to excretion in the bile. These results explain the beneficial effects of probucol on atherosclerosis and xanthomas despite its HDL-lowering effects and suggest that inactivation of hepatic ABCA1 leads to increased RCT despite reducing plasma HDL-C levels.
Tumorigenicity is an associated risk for transplantation of hepatocytes differentiated from human induced pluripotent stem (hiPS) cells. Hepatocytes express the enzymes galactokinase and ornithine ...transcarbamylase (OTC) to aid in their own survival. However, hiPS cells do not express these enzymes, and therefore, are not be expected to survive in a medium containing galactose and ornithine and lacking glucose and arginine.
Real-time quantitative polymerase chain reaction (PCR) was performed to analyze the expression of galactokinase 1 (GALK1)1 and GALK2, ornithine carbamyltransferase, and phenylalanine hydroxylase (PAH). The hiPS cell line 201B7 was cultured in hepatocyte selection medium (HSM), which lacks glucose and arginine but contains galactose and ornithine. Furthermore, microscopic analysis of the cultured cells was performed after hematoxylin and eosin (H&E) staining, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL). The hiPS cells were immunostained to assess their pluripotency in HSM. In addition, the primary human hepatocytes were cultured with or without hiPS cells in HSM.
The expression levels of GALK1, GALK2, OTC, and PAH in 201B7 were 22.2±5.0 (average ± standard deviation), 14.2% ±1.1%, 1.2% ±0.2%, and 8.4% ±0.7% respectively, compared with those in the adult liver. The hiPS cell population diminished when cultured in HSM and completely disappeared after 3 days. The cultured cells showed condensation or fragmentation of their nuclei, thereby suggesting apoptosis. TUNEL staining confirmed that the cells had undergone apoptosis. The 201B7 cells were positive for Nanog, SSEA-4, and TRA-1-60. The primary human hepatocytes survived when cultured alone in HSM and when co-cultured with hiPS cells.
Therefore, HSM is and ideal medium for eliminating hiPS cells and purifying hepatocytes without inducing any damage.
Hepatocyte differentiation inducer (HDI) lacks both glucose and arginine, but is supplemented with galactose and ornithine, and is added together with other reagents such as apoptosis inhibitor and ...oncostatin M. Although human induced pluripotent stem (iPS) cells initiate hepatocyte differentiation, most die within 7 days. In this study, we investigated both HDI and conventional media for their potential to improve cell survival.
201B7 iPS cells were cultured in conventional media. This consisted of three cycles of 5-day culture in William's E (WE) medium, followed by a 2-day culture in HDI.
Expression levels of α-feto protein (AFP) were higher in cells cultured in WE and in Dulbecco's Modified Eagle's Medium/Nutrient F-12 Ham (DF12). 201B7 cells expressed the highest AFP and albumin (ALB) when cultured in HDI for 2 days following 7-day culture in WE. After three cycles of 5-day culture in WE followed by 2 days in HDI, 201B7 cells expressed AFP and ALB 54 ± 2.3 (average ± standard deviation) and 73 ± 15.1 times higher, respectively, than those cultured in ReproFF (feeder-free condition).
201B7 cells survived culture in WE for 7 days followed HDI for 2 days. After three cycles of culture under these conditions, hepatocyte differentiation was enhanced, as evidenced by increased AFP and ALB expression.
AIM:To develop a molecular therapy for pancreatic cancer, the insulin-like growth factor-I (IGF-I) signaling pathway was analyzed.METHODS: Pancreatic cancer cell lines (MIA-Paca2, NOR-P1, PANC-1, ...PK-45H, PK-1, PK-59 and KP-4) were cultured in media with 10 mL/L fetal bovine serum. Western blotting analysis was performed to clarify the expression of IGF-I receptor (IGF-IR). Picropodophyllin (PPP), a specific inhibitor of IGF-IR, LY294002, a specific inhibitor of phosphatidylinositol3 kinase (PI3K), and PD980...
AIM: To investigate the early upper gastrointestinal endoscopy(endoscopy) significantly reduces mortality resulting from upper gastrointestinal(GI) bleeding. METHODS: Upper GI bleeding was defined as ...1a, 1b, 2a, and 2b according to the Forrest classification. The hemoglobin(Hb), and C-reactive protein(CRP) were examined at around the day of endoscopy and 3 mo prior to endoscopy. The rate of change was calculated as follows:(the result of blood examination on the day of endoscopy- the results of blood examination 3 mo prior to endoscopy)/(results of blood examination 3 mo prior to endoscopy). Receiver operating characteristic curves were created to determine threshold values. RESULTS: Seventy-nine men and 77 women were enrolled. There were 17 patients with upper GI bleeding: 12 with a gastric ulcer, 3 with a duodenal ulcer, 1 with an acute gastric mucosal lesion, and 1 with gastric cancer. The area under the curve(AUC), threshold, sensitivity, and specificity of Hb around the day of endoscopy were 0.902, 11.7 g/dL, 94.1%, and 77.1%, respectively, while those of CRP were 0.722, 0.5 mg/dL, 70.5%, and 73%, respectively. The AUC, threshold, sensitivity, and specificity of the rate of change of Hb were 0.851,-21.3%, 76.4%, and 82.6%, respectively, while those of CRP were 0.901, 100%, 100%, and 82.5%, respectively. CONCLUSION: Predictors for upper GI bleeding were Hb < 11.7 g/dL, reduction rate in the Hb > 21.3% and an increase in the CRP > 100%, 3 mo before endoscopy.
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