A reliable and sensitive complex I assay is an essential tool for the diagnosis of mitochondrial disorders, but current spectrophotometric assays suffer from low sensitivity, low specificity, or ...both. This deficiency is mainly due to the poor solubility of coenzyme-Q analogs and reaction mixture turbidity caused by the relatively high concentrations of tissue extract that are often required to measure complex I.
We developed a new spectrophotometric assay to measure complex I in mitochondrial fractions and applied it to muscle and cultured fibroblasts. The method is based on measuring 2,6-dichloroindophenol reduction by electrons accepted from decylubiquinol, reduced after oxidation of NADH by complex I. The assay thus is designed to avoid nonspecific NADH oxidation because electrons produced in these reactions are not accepted by decylubiquinone, resulting in high rotenone sensitivity.
The assay was linear with time and amount of mitochondria. The K(m) values for NADH and 2,6-dichloroindophenol in muscle mitochondria were 0.04 and 0.017 mmol/L, respectively. The highest complex I activities were measured with 0.07 mmol/L decylubiquinone and 3.5 g/L bovine serum albumin. The latter was an essential component of the reaction mixture, increasing the solubility of decylubiquinone and rotenone. In patients with previously diagnosed complex I deficiencies, the new assay detected the complex I deficiencies in both muscle and fibroblasts.
This spectrophotometric assay is reproducible, sensitive, and specific for complex I activity because of its high rotenone sensitivity, and it can be applied successfully to the diagnosis of complex I deficiencies.
In renal transplantation, use of calcineurin inhibitors (CNIs) is associated with nephrotoxicity and immunosuppression with malignancies and infections. This trial aimed to minimize CNI exposure and ...total immunosuppression while maintaining efficacy. We performed a randomized controlled, open‐label multicenter trial with early cyclosporine A (CsA) elimination. Patients started with basiliximab, prednisolone (P), mycophenolate sodium (MPS), and CsA. At 6 months, immunosuppression was tapered to P/CsA, P/MPS, or P/everolimus (EVL). Primary outcomes were renal fibrosis and inflammation. Secondary outcomes were estimated glomerular filtration rate (eGFR) and incidence of rejection at 24 months. The P/MPS arm was prematurely halted. The trial continued with P/CsA (N = 89) and P/EVL (N = 96). Interstitial fibrosis and inflammation were significantly decreased and the eGFR was significantly higher in the P/EVL arm. Cumulative rejection rates were 13% (P/EVL) and 19% (P/CsA), (p = 0.08). A post hoc analysis of HLA and donor‐specific antibodies at 1 year after transplantation revealed no differences. An individualized immunosuppressive strategy of early CNI elimination to dual therapy with everolimus was associated with decreased allograft fibrosis, preserved allograft function, and good efficacy, but also with more serious adverse events and discontinuation. This can be a valuable alternative regimen in patients suffering from CNI toxicity.
This randomized trial in renal transplant recipients shows that with tight monitoring of drug levels, a triple calcineurin inhibitor–containing drug maintenance regimen can be tapered to a double calcineurin‐free drug regimen, and that this regimen is associated with better renal function and less interstitial fibrosis and inflammation.
Intervening in angiotensin (Ang)-II type 2 receptor (AT2) signaling may have therapeutic potential for bronchopulmonary dysplasia (BPD) by attenuating lung inflammation and preventing arterial ...hypertension (PAH)-induced right ventricular hypertrophy (RVH). We first investigated the role of AT2 inhibition with PD123319 (0.5 and 2 mg·kg(-1)·day(-1)) on the beneficial effect of AT2 agonist LP2-3 (5 μg/kg twice a day) on RVH in newborn rats with hyperoxia-induced BPD. Next we determined the cardiopulmonary effects of PD123319 (0.1 mg·kg(-1)·day(-1)) in two models: early treatment during continuous exposure to hyperoxia for 10 days and late treatment starting on day 6 in rat pups exposed postnatally to hyperoxia for 9 days, followed by a 9-day recovery period in room air. Parameters investigated included lung and heart histopathology, fibrin deposition, vascular leakage, and differential mRNA expression. Ten days of coadministration of LP2-3 and PD123319 abolished the beneficial effects of LP2-3 on RVH in experimental BPD. In the early treatment model PD123319 attenuated cardiopulmonary injury by reducing alveolar septal thickness, pulmonary influx of inflammatory cells, including macrophages and neutrophils, medial wall thickness of small arterioles, and extravascular collagen III deposition, and by preventing RVH. In the late treatment model PD123319 diminished PAH and RVH, demonstrating that PAH is reversible in the neonatal period. At high concentrations PD123319 blocks the beneficial effects of the AT2-agonist LP2-3 on RVH. At low concentrations PD123319 attenuates cardiopulmonary injury by reducing pulmonary inflammation and fibrosis and preventing PAH-induced RVH but does not affect alveolar and vascular development in newborn rats with experimental BPD.
Non-local fluctuation phenomena in liquids Croccolo, F.; Ortiz de Zárate, J. M.; Sengers, J. V.
The European physical journal. E, Soft matter and biological physics,
12/2016, Letnik:
39, Številka:
12
Journal Article
Recenzirano
.
Fluids in non-equilibrium steady states exhibit long-range fluctuations which extend over the entire system. They can be described by non-equilibrium thermodynamics and fluctuating hydrodynamics ...that assume local equilibrium for the thermophysical properties as a function of space and time. The experimental evidence for the consistency between this assumption of local equilibrium in the equations and the non-local fluctuation phenomena observed is reviewed.
Graphical abstract
Insufficient hemodynamics during agonal phase—ie, the period between withdrawal of life‐sustaining treatment and circulatory arrest—in Maastricht category III circulatory‐death donors (DCD) ...potentially exacerbate ischemia/reperfusion injury. We included 409 Dutch adult recipients of DCD donor kidneys transplanted between 2006 and 2014. Peripheral oxygen saturation (SpO2‐with pulse oximetry at the fingertip) and systolic blood pressure (SBP‐with arterial catheter) were measured during agonal phase, and were dichotomized into minutes of SpO2 > 60% or SpO2 < 60%, and minutes of SBP > 80 mmHg or SBP < 80 mmHg. Outcome measures were and primary non‐function (PNF), delayed graft function (DGF), and three‐year graft survival. Primary non‐function (PNF) rate was 6.6%, delayed graft function (DGF) rate was 67%, and graft survival at three years was 76%. Longer periods of agonal phase (median 16 min IQR 11‐23) contributed significantly to an increased risk of DGF (P = .012), but not to PNF (P = .071) and graft failure (P = .528). Multiple logistic regression analysis showed that an increase from 7 to 20 minutes in period of SBP < 80 mmHg was associated with 2.19 times the odds (95% CI 1.08‐4.46, P = .030) for DGF. In conclusion, duration of agonal phase is associated with early transplant outcome. SBP < 80 mmHg during agonal phase shows a better discrimination for transplant outcome than SpO2 < 60% does.
This investigation of the predictive ability of hemodynamics during the agonal phase of circulatory death donors shows that systolic blood pressure discriminates short‐term transplant outcomes better than peripheral oxygenation does.
A systematic review of qualitative and quantitative studies.
To summarize evidence from studies among patients with low back pain on their expectations and satisfaction with treatment as part of ...practice guideline development.
Patients are often dissatisfied with treatment for acute or chronic back pain.
We searched the literature for studies on patient expectations and satisfaction with treatment for low back pain. Treatment aspects related to expectations or satisfaction were identified in qualitative studies. Percentages of dissatisfied patients were calculated from quantitative studies.
Twelve qualitative and eight quantitative studies were found. Qualitative studies revealed the following aspects that patient expectation from treatment for back pain or with which they are dissatisfied. Patients want a clear diagnosis of the cause of their pain, information and instructions, pain relief, and a physical examination. Next, expectations are that there are more diagnostic tests, other therapy or referrals to specialists, and sickness certification. They expect confirmation from the healthcare provider that their pain is real. Like other patients, they want a confidence-based association that includes understanding, listening, respect, and being included in decision-making. The results from qualitative studies are confirmed by quantitative studies.
Patients have explicit expectations on diagnosis, instructions, and interpersonal management. New strategies need to be developed in order to meet patients' expectations better. Practice guidelines should pay more attention to the best way of discussing the causes and diagnosis with the patient and should involve them in the decision-making process.
Diagnosis of mitochondrial disorders usually requires a muscle biopsy to examine mitochondrial function. We describe our diagnostic procedure and results for 29 patients with mitochondrial disorders.
...Muscle biopsies were from 43 healthy individuals and 29 patients with defects in one of the oxidative phosphorylation (OXPHOS) complexes, the pyruvate dehydrogenase complex (PDHc), or the adenine nucleotide translocator (ANT). Homogenized muscle samples were used to determine the oxidation rates of radiolabeled pyruvate, malate, and succinate in the absence or presence of various acetyl Co-A donors and acceptors, as well as specific inhibitors of tricarboxylic acid cycle or OXPHOS enzymes. We determined the rate of ATP production from oxidation of pyruvate.
Each defect in the energy-generating system produced a specific combination of substrate oxidation impairments. PDHc deficiencies decreased substrate oxidation reactions containing pyruvate. Defects in complexes I, III, and IV decreased oxidation of pyruvate plus malate, with normal to mildly diminished oxidation of pyruvate plus carnitine. In complex V defects, pyruvate oxidation improved by addition of carbonyl cyanide 3-chlorophenyl hydrazone, whereas other oxidation rates were decreased. In most patients, ATP production was decreased.
The proposed method can be successfully applied to the diagnosis of defects in PDHc, OXPHOS complexes, and ANT.
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