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Objective: Storage of triglycerides in lipid droplets is governed by a set of lipid droplet-associated proteins. One of these lipid droplet-associated proteins, hypoxia-inducible lipid droplet-associated (HILPDA), was found to impair lipid droplet breakdown in macrophages and cancer cells by inhibiting adipose triglyceride lipase. Here, we aimed to better characterize the role and mechanism of action of HILPDA in hepatocytes. Methods: We performed studies in HILPDA-deficient and HILPDA-overexpressing liver cells, liver slices, and mice. The functional role and physical interactions of HILPDA were investigated using a variety of biochemical and microscopic techniques, including real-time fluorescence live-cell imaging and Förster resonance energy transfer-fluorescence lifetime imaging microscopy (FRET-FLIM). Results: Levels of HILPDA were markedly induced by fatty acids in several hepatoma cell lines. Hepatocyte-specific deficiency of HILPDA in mice modestly but significantly reduced hepatic triglycerides in mice with non-alcoholic steatohepatitis. Similarly, deficiency of HILPDA in mouse liver slices and primary hepatocytes reduced lipid storage and accumulation of fluorescently-labeled fatty acids in lipid droplets, respectively, which was independent of adipose triglyceride lipase. Fluorescence microscopy showed that HILPDA partly colocalizes with lipid droplets and with the endoplasmic reticulum, is especially abundant in perinuclear areas, and mainly associates with newly added fatty acids. Real-time fluorescence live-cell imaging further revealed that HILPDA preferentially localizes to lipid droplets that are being remodeled. Overexpression of HILPDA in liver cells increased the activity of diacylglycerol acyltransferases (DGAT) and DGAT1 protein levels, concurrent with increased lipid storage. Confocal microscopy coupled to FRET-FLIM analysis demonstrated that HILPDA physically interacts with DGAT1 in living liver cells. The stimulatory effect of HILPDA on lip

Mild cold acclimation for 10 days has been previously shown to markedly improve insulin sensitivity in patients with type 2 diabetes. Here we show in a single-arm intervention study (Trialregister.nl ID: NL4469/NTR5711) in nine patients with type 2 diabetes that ten days of mild cold acclimation (16–17 °C) in which observable, overt shivering was prevented, does not result in improved insulin sensitivity, postprandial glucose and lipid metabolism or intrahepatic lipid content and only results in mild effects on overnight fasted fat oxidation, postprandial energy expenditure and aortic augmentation index. The lack of marked metabolic effects in this study is associated with a lack of self-reported shivering and a lack of upregulation of gene expression of muscle activation or muscle contraction pathways in skeletal muscle and suggests that some form of muscle contraction is needed for beneficial effects of mild cold acclimation.

Objective: Intramyocellular lipid droplets (LD) and their coat proteins PLIN2 and PLIN5 are involved in lipolysis, with a putative role for PLIN5 in mitochondrial tethering. Reportedly, these proteins co-localize and cover the surface of the LD. To provide the spatial basis for understanding how these proteins possess their distinct roles, we examined the precise location of PLIN2 and PLIN5 and explored PLIN5 presence at LD-mitochondria contact sites using Stimulated emission depletion (STED) microscopy and correlative light-electron microscopy (CLEM) in human skeletal muscle sections.Methods: LDs were stained by MDH together with combinations of mitochondria] proteins and PLINs. Subcellular distribution and co-localization of PLIN proteins and mitochondria was imaged by STED microscopy (Leica TCS SP8) and quantified using Pearson's correlation coefficients and intensity profile plots. CLEM was employed to examine the presence of PLIN5 on mitochondria-LD contact sites.Results: Both PLIN2 and PLIN5 localized to the LD in a dot-like, juxtaposed fashion rather than colocalizing and covering the entire LD. Both STED and CLEM revealed a high fraction of PLIN5 at the LD-mitochondria interface, but not at mitochondrial cristae, as suggested previously.Conclusion: Using two super-resolution imaging approaches, this is the first study to show that in sections of human skeletal muscle PLIN2 and PLIN5 localize to the LD at distinct sites, with abundance of PLIN5 at LD-mitochondria tethering sites. This novel spatial information uncovers that PLIN proteins do not serve as lipolytic barriers but rather are docking sites for proteins facilitating selective lipase access under a variety of lipolytic conditions.

Intramyocellular lipid (IMCL) hampers insulin sensitivity, albeit not in endurance-trained athletes (Trained). Compared to type 2 diabetes mellitus (T2DM) patients, Trained subjects have high levels of perilipin 5 (PLIN5). In the present study, we tested whether the fraction of PLIN5-coated lipid droplets (LDs) is a determinant of skeletal muscle insulin sensitivity and contributes to the athlete's paradox. Muscle biopsies were taken from eight Trained, Lean sedentary, Obese and T2DM subjects. Trained, Obese and T2DM subjects were matched for total IMCL content. Confocal images were analysed for lipid area fraction, LD size and number and PLIN5+ and PLIN5- LDs were measured. A stepwise linear regression was performed to identify factors explaining observed variance in glucose infusion rate (GIR). Trained and T2DM subjects stored IMCL differently; Trained subjects had a higher number of LDs compared to T2DM subjects (0.0370.004m(-2)vs. 0.0230.003m(-2), P=0.024) that were non-significantly smaller (0.270.01m(2)vs. 0.320.02m(2), P=0.197, Trained vs. T2DM). Even though total PLIN5 protein content was almost double in Trained vs. T2DM subjects (1.650.21AU vs. 0.89 +/- 0.09AU, P=0.004), PLIN5 coating did not affect LD number or size significantly. Of the observed variance in GIR, the largest fraction by far (70.2%) was explained by maximal oxygen uptake. Adding PLIN5 protein content or PLIN5+ LDs increased the explained variance in GIR (74.7% and 80.7% for PLIN5 protein content and PLIN5+ LDs, respectively). Thus, the putative relationship between PLIN5 and insulin sensitivity is at best indirect and is apparent only in conjunction with maximal oxygen uptake. Hence, PLIN5 abundance cannot be causally linked to the athlete's paradox.

01 juni 2020, Contains fulltext : 220950.pdf (Publisher’s version ) (Open Access), AIMS/HYPOTHESIS: Physical inactivity, low mitochondrial function, increased intramyocellular lipid (IMCL) deposition and reduced insulin sensitivity are common denominators of chronic metabolic disorders, like obesity and type 2 diabetes. Yet, whether low mitochondrial function predisposes to insulin resistance in humans is still unknown. METHODS: Here we investigated, in an intervention study, whether muscle with low mitochondrial oxidative capacity, induced by one-legged physical inactivity, would feature stronger signs of lipid-induced insulin resistance. To this end, ten male participants (age 22.4 ± 4.2 years, BMI 21.3 ± 2.0 kg/m(2)) underwent a 12 day unilateral lower-limb suspension with the contralateral leg serving as an active internal control. RESULTS: In vivo, mitochondrial oxidative capacity, assessed by phosphocreatine (PCr)-recovery half-time, was lower in the inactive vs active leg. Ex vivo, palmitate oxidation to (14)CO(2) was lower in the suspended leg vs the active leg; however, this did not result in significantly higher [(14)C]palmitate incorporation into triacylglycerol. The reduced mitochondrial function in the suspended leg was, however, paralleled by augmented IMCL content in both musculus tibialis anterior and musculus vastus lateralis, and by increased membrane bound protein kinase C (PKC) θ. Finally, upon lipid infusion, insulin signalling was lower in the suspended vs active leg. CONCLUSIONS/INTERPRETATION: Together, these results demonstrate, in a unique human in vivo model, that a low mitochondrial oxidative capacity due to physical inactivity directly impacts IMCL accumulation and PKCθ translocation, resulting in impaired insulin signalling upon lipid infusion. This demonstrates the importance of mitochondrial oxidative capacity and muscle fat accumulation in the development of insulin resistance in humans. TRIAL REGISTRATION: ClinicalTrial.gov NCT01576250. FUNDING: PS was supported by a 'VICI' Research Grant for innovative research f

Skeletal muscle can store excess fat as subcellular lipid droplets (LDs). While originally viewed as uninteresting static balls of triacylglycerol, it is now clear that myocellular LDs play an active role in myocellular (patho) physiology. In this review we aim to discuss the role of LDs in muscle cell insulin sensitivity and identify parameters which appear to affect this relationship. Moreover, we discuss the application of novel tools permitting detailed examination of these parameters. This article is part of a Special Issue entitled: Recent Advances in Lipid Droplet Biology edited by Rosalind Coleman and Matthijs Hesselink.

Insulin resistance lies at the basis of type 2 diabetes. Insulin resistance is often paired with increased fat deposits in muscle tissue, which some people claim to be the cause; however, highly trained and insulin-sensitive athletes also store a lot of fat in their muscles, which serves to counter this causal relationship. This dissertation examines the distinct characteristics of fat accumulation in insulin-sensitive and insulin-resistant states using techniques such as microscopy. The most important conclusion is that normal lipid droplets that come into contact with the protein PLIN5 are associated with higher insulin sensitivity.

Exercise training reduces intrahepatic lipid (IHL) content in people with elevated liver fat content. It is unclear, however, whether exercise training reduces IHL content in people with normal liver fat content. Here, we measured the effect of exercise training on IHL content in people with and people without nonalcohol fatty liver. We further measured changes in insulin sensitivity and hepatic energy metabolism. Eleven males with nonalcoholic fatty liver (NAFL) and 11 body mass index-matched individuals without nonalcoholic fatty liver (CON) completed a 12-wk supervised exercise training program. IHL content (proton magnetic resonance spectroscopy), maximal oxidative capacity (O(2)max, spiroergometry), total muscle strength, body composition, insulin sensitivity (hyperinsulinemic-euglycemic clamp), hepatic ATP-to-total phosphorus ratio, and the hepatic phosphomonoester-to-phosphodiester (PME/PDE) ratio (phosphorus magnetic resonance spectroscopy) were determined. IHL content reduced with exercise training (P = 0.014) in the whole study population. The relative reduction in IHL content was comparable in NAFL (-34.5 +/- 54.0%) and CON (-28.3 +/- 60.1%) individuals (P = 0.800). O(2)max (P <0.001), total muscle strength (P <0.001), and skeletal muscle insulin sensitivity (P = 0.004) increased, whereas adipose tissue (P = 0.246) and hepatic (P = 0.086) insulin sensitivity did not increase significantly. Hepatic ATP-to-total phosphorus ratio (P = 0.987) and PME/PDE ratio (P = 0.792) did not change. Changes in IHL content correlated with changes in body weight (r = 0.451, P = 0.035) and changes in hepatic PME/PDE ratio (r = 0.569, P = 0.019). In conclusion, exercise training reduced intrahepatic lipid content in people with nonalcoholic fatty liver and in people with normal intrahepatic lipid content, and the percent reduction in intrahepatic lipid content was similar in both groups.

Intramyocellular lipid (IMCL) hampers insulin sensitivity, albeit not in endurance-trained athletes (Trained). Compared to type 2 diabetes mellitus (T2DM) patients, Trained subjects have high levels of perilipin 5 (PLIN5). In the present study, we tested whether the fraction of PLIN5-coated lipid droplets (LDs) is a determinant of skeletal muscle insulin sensitivity and contributes to the athlete's paradox. Muscle biopsies were taken from eight Trained, Lean sedentary, Obese and T2DM subjects. Trained, Obese and T2DM subjects were matched for total IMCL content. Confocal images were analysed for lipid area fraction, LD size and number and PLIN5+ and PLIN5- LDs were measured. A stepwise linear regression was performed to identify factors explaining observed variance in glucose infusion rate (GIR). Trained and T2DM subjects stored IMCL differently; Trained subjects had a higher number of LDs compared to T2DM subjects (0.0370.004m(-2)vs. 0.0230.003m(-2), P=0.024) that were non-significantly smaller (0.270.01m(2)vs. 0.320.02m(2), P=0.197, Trained vs. T2DM). Even though total PLIN5 protein content was almost double in Trained vs. T2DM subjects (1.650.21AU vs. 0.89 +/- 0.09AU, P=0.004), PLIN5 coating did not affect LD number or size significantly. Of the observed variance in GIR, the largest fraction by far (70.2%) was explained by maximal oxygen uptake. Adding PLIN5 protein content or PLIN5+ LDs increased the explained variance in GIR (74.7% and 80.7% for PLIN5 protein content and PLIN5+ LDs, respectively). Thus, the putative relationship between PLIN5 and insulin sensitivity is at best indirect and is apparent only in conjunction with maximal oxygen uptake. Hence, PLIN5 abundance cannot be causally linked to the athlete's paradox.

Objective: Intramyocellular lipid droplets (LD) and their coat proteins PLIN2 and PLIN5 are involved in lipolysis, with a putative role for PLIN5 in mitochondrial tethering. Reportedly, these proteins co-localize and cover the surface of the LD. To provide the spatial basis for understanding how these proteins possess their distinct roles, we examined the precise location of PLIN2 and PLIN5 and explored PLIN5 presence at LD-mitochondria contact sites using Stimulated emission depletion (STED) microscopy and correlative light-electron microscopy (CLEM) in human skeletal muscle sections.Methods: LDs were stained by MDH together with combinations of mitochondria] proteins and PLINs. Subcellular distribution and co-localization of PLIN proteins and mitochondria was imaged by STED microscopy (Leica TCS SP8) and quantified using Pearson's correlation coefficients and intensity profile plots. CLEM was employed to examine the presence of PLIN5 on mitochondria-LD contact sites.Results: Both PLIN2 and PLIN5 localized to the LD in a dot-like, juxtaposed fashion rather than colocalizing and covering the entire LD. Both STED and CLEM revealed a high fraction of PLIN5 at the LD-mitochondria interface, but not at mitochondrial cristae, as suggested previously.Conclusion: Using two super-resolution imaging approaches, this is the first study to show that in sections of human skeletal muscle PLIN2 and PLIN5 localize to the LD at distinct sites, with abundance of PLIN5 at LD-mitochondria tethering sites. This novel spatial information uncovers that PLIN proteins do not serve as lipolytic barriers but rather are docking sites for proteins facilitating selective lipase access under a variety of lipolytic conditions.

Exercise training reduces intrahepatic lipid (IHL) content in people with elevated liver fat content. It is unclear, however, whether exercise training reduces IHL content in people with normal liver fat content. Here, we measured the effect of exercise training on IHL content in people with and people without nonalcohol fatty liver. We further measured changes in insulin sensitivity and hepatic energy metabolism. Eleven males with nonalcoholic fatty liver (NAFL) and 11 body mass index-matched individuals without nonalcoholic fatty liver (CON) completed a 12-wk supervised exercise training program. IHL content (proton magnetic resonance spectroscopy), maximal oxidative capacity (O(2)max, spiroergometry), total muscle strength, body composition, insulin sensitivity (hyperinsulinemic-euglycemic clamp), hepatic ATP-to-total phosphorus ratio, and the hepatic phosphomonoester-to-phosphodiester (PME/PDE) ratio (phosphorus magnetic resonance spectroscopy) were determined. IHL content reduced with exercise training (P = 0.014) in the whole study population. The relative reduction in IHL content was comparable in NAFL (-34.5 +/- 54.0%) and CON (-28.3 +/- 60.1%) individuals (P = 0.800). O(2)max (P <0.001), total muscle strength (P <0.001), and skeletal muscle insulin sensitivity (P = 0.004) increased, whereas adipose tissue (P = 0.246) and hepatic (P = 0.086) insulin sensitivity did not increase significantly. Hepatic ATP-to-total phosphorus ratio (P = 0.987) and PME/PDE ratio (P = 0.792) did not change. Changes in IHL content correlated with changes in body weight (r = 0.451, P = 0.035) and changes in hepatic PME/PDE ratio (r = 0.569, P = 0.019). In conclusion, exercise training reduced intrahepatic lipid content in people with nonalcoholic fatty liver and in people with normal intrahepatic lipid content, and the percent reduction in intrahepatic lipid content was similar in both groups.

In recent years digitalization and other global trends have resulted in a change in the way that maintenance is performed in many industries. There is a large body of research on the subject of maintenance policy in general, and an ever-growing wealth of technical studies that detail new maintenance techniques. However, the majority of maintenance research focuses on high risk sectors, closed systems, or is highly application oriented. Very little research can be found on maintenance decision making and policy choices for less controlled or less straightforward maintenance applications, such as the maintenance of escalators dispersed throughout a building or a city. This research builds on the existing knowledge and literature about maintenance policy options and how they are implemented in various industries and investigates maintenance policy decision making in the context of transfer systems. It introduces a common set of terminology around maintenance policy choices that should improve clarity and communication. Matrices have been developed to classify transfer system types based on their system structure and flow characteristics. System types have been named and described as light, stable, dynamic and heavy loads based on their features, and four maintenance policies have been defined and matched to the system types based on the literature and empirical findings. This work explains why digitalized, condition-based maintenance should be considered as two separate categories of maintenance depending on whether the primary mode of maintenance decision making is time-based or condition-based. This work provides clarity into what outcomes, benefits and risks can be expected from different kinds of maintenance. The primary motivating factor in this research is to improve escalator safety. While it is known that better maintenance practices result in safer escalators, the lack of research in this area means that the root causes of escalator accidents often aren’t clea

Non-alcoholic fatty liver (NAFL) is an independent risk factor for the development of type 2 diabetes (T2DM). We examined metabolic perturbations in patients with NAFL, patients with T2DM, and control (CON) subjects with normal intrahepatic lipid (IHL) content. A two-step (10 mU/m(2) /min; 40 mU/m(2)/min) hyperinsulinemic-euglycemic clamp was performed in 11 NAFL, 13 T2DM, and 11 CON subjects, all matched for BMI, and aerobic fitness. IHL content was measured using proton magnetic resonance spectroscopy. Because of high IHL content variability in T2DM patients, this group was separated into a high IHL content group (IHL >= 5.0%, T2DM+ NAFL) and a normal IHL content group (IHL <5.0%, T2DM-non-NAFL) for further analysis.IHL content was increased in NAFL and T2DM+ NAFL subjects (P <0.050 versus CON and T2DM-non-NAFL subjects). Adipose tissue insulin sensitivity index (Adipo-IRi) was higher in NAFL (P<0.050 versus CON and T2DM-non-NAFL subjects) and in T2DM+ NAFL subjects (P= 0.055 versus CON subjects, P<0.050 versus T2DM-non-NAFL subjects). Suppression of plasma-free fatty acids (P= 0.046) was lower in NAFL compared with CON subjects, with intermediate values for T2DM-non-NAFL, and T2DM+ NAFL subjects. Suppression of endogenous glucose production (EGP) and insulin-stimulated glucose disposal (Delta R-d) was comparable between NAFL, T2DM-non-NAFL, and T2DM+ NAFL subjects (all P> 0.05), and was lower in comparison with CON subjects (all P<0.01). Metabolic flexibility was lower in T2DM-non-NAFL subjects (P= 0.047) and NAFL subjects (P= 0.059) compared with CON subjects. Adipo-IRi (r= 0.652, P<0.001), hepatic insulin resistance index (HIRi) (r= 0.576, P= 0.001), and Delta R-d (r=-0.653, P<0.001) correlated with IHL content.Individuals with NAFL suffer from metabolic perturbations to a similar degree as T2DM patients. NAFL is an important feature leading to severe insulin resistance and should be viewed as a serious health threat

Non-alcoholic fatty liver (NAFL) is an independent risk factor for the development of type 2 diabetes (T2DM). We examined metabolic perturbations in patients with NAFL, patients with T2DM, and control (CON) subjects with normal intrahepatic lipid (IHL) content. A two-step (10 mU/m(2) /min; 40 mU/m(2)/min) hyperinsulinemic-euglycemic clamp was performed in 11 NAFL, 13 T2DM, and 11 CON subjects, all matched for BMI, and aerobic fitness. IHL content was measured using proton magnetic resonance spectroscopy. Because of high IHL content variability in T2DM patients, this group was separated into a high IHL content group (IHL >= 5.0%, T2DM+ NAFL) and a normal IHL content group (IHL <5.0%, T2DM-non-NAFL) for further analysis.IHL content was increased in NAFL and T2DM+ NAFL subjects (P <0.050 versus CON and T2DM-non-NAFL subjects). Adipose tissue insulin sensitivity index (Adipo-IRi) was higher in NAFL (P<0.050 versus CON and T2DM-non-NAFL subjects) and in T2DM+ NAFL subjects (P= 0.055 versus CON subjects, P<0.050 versus T2DM-non-NAFL subjects). Suppression of plasma-free fatty acids (P= 0.046) was lower in NAFL compared with CON subjects, with intermediate values for T2DM-non-NAFL, and T2DM+ NAFL subjects. Suppression of endogenous glucose production (EGP) and insulin-stimulated glucose disposal (Delta R-d) was comparable between NAFL, T2DM-non-NAFL, and T2DM+ NAFL subjects (all P> 0.05), and was lower in comparison with CON subjects (all P<0.01). Metabolic flexibility was lower in T2DM-non-NAFL subjects (P= 0.047) and NAFL subjects (P= 0.059) compared with CON subjects. Adipo-IRi (r= 0.652, P<0.001), hepatic insulin resistance index (HIRi) (r= 0.576, P= 0.001), and Delta R-d (r=-0.653, P<0.001) correlated with IHL content.Individuals with NAFL suffer from metabolic perturbations to a similar degree as T2DM patients. NAFL is an important feature leading to severe insulin resistance and should be viewed as a serious health threat

Skeletal muscle can store excess fat as subcellular lipid droplets (LDs). While originally viewed as uninteresting static balls of triacylglycerol, it is now clear that myocellular LDs play an active role in myocellular (patho) physiology. In this review we aim to discuss the role of LDs in muscle cell insulin sensitivity and identify parameters which appear to affect this relationship. Moreover, we discuss the application of novel tools permitting detailed examination of these parameters. This article is part of a Special Issue entitled: Recent Advances in Lipid Droplet Biology edited by Rosalind Coleman and Matthijs Hesselink.

Insulin resistance lies at the basis of type 2 diabetes. Insulin resistance is often paired with increased fat deposits in muscle tissue, which some people claim to be the cause; however, highly trained and insulin-sensitive athletes also store a lot of fat in their muscles, which serves to counter this causal relationship. This dissertation examines the distinct characteristics of fat accumulation in insulin-sensitive and insulin-resistant states using techniques such as microscopy. The most important conclusion is that normal lipid droplets that come into contact with the protein PLIN5 are associated with higher insulin sensitivity.

Item does not contain fulltext, OBJECTIVE: Parental high-fat feeding was proposed to negatively impact metabolic health in offspring. Here, the ectopic fat storage in heart and liver in offspring was investigated, and the effects on mitochondrial function, de novo lipogenesis, and postprandial lipid metabolism were explored in detail. METHODS: Male and female mice received either a high-fat (HF) or standard chow (LF) diet during mating, gestation and lactation. All offspring animals received the HF diet. RESULTS: Abdominal visceral adipose tissue tended to be higher in HF/HF mice. Cardiac lipid content was also higher in the HF/HF mice (LF/HF vs. HF/HF: 1.03% +/- 0.08% vs. 1.33% +/- 0.07% of water signal, P = 0.01). In contrast, hepatic lipid content tended to be lower in HF/HF mice compared to LF/HF mice. A severely disturbed postprandial lipid clearance was revealed in HF/HF mice by the results from the triglyceride (TG) tolerance tests (LF/HF vs. HF/HF: 6,753 +/- 2,213 vs. 14,367 +/- 1,978 mmol l(-1) min(-1) , P = 0.01) and (13) C-fatty acid retention test (LF/HF vs. HF/HF: 2.73% +/- 0.85% vs. 0.89% +/- 0.26% retention from bolus, P = 0.04), which may underlie the lower hepatic lipid content. CONCLUSIONS: Here it is shown that HF diet negatively impacts postprandial TG clearance in offspring and results in an overall metabolic unfavorable phenotype and ectopic lipid deposition in the heart and in visceral storage sites.

OBJECTIVE: Parental high-fat feeding was proposed to negatively impact metabolic health in offspring. Here, the ectopic fat storage in heart and liver in offspring was investigated, and the effects on mitochondrial function, de novo lipogenesis, and postprandial lipid metabolism were explored in detail. METHODS: Male and female mice received either a high-fat (HF) or standard chow (LF) diet during mating, gestation and lactation. All offspring animals received the HF diet. RESULTS: Abdominal visceral adipose tissue tended to be higher in HF/HF mice. Cardiac lipid content was also higher in the HF/HF mice (LF/HF vs. HF/HF: 1.03% +/- 0.08% vs. 1.33% +/- 0.07% of water signal, P = 0.01). In contrast, hepatic lipid content tended to be lower in HF/HF mice compared to LF/HF mice. A severely disturbed postprandial lipid clearance was revealed in HF/HF mice by the results from the triglyceride (TG) tolerance tests (LF/HF vs. HF/HF: 6,753 +/- 2,213 vs. 14,367 +/- 1,978 mmol l(-1) min(-1) , P = 0.01) and (13) C-fatty acid retention test (LF/HF vs. HF/HF: 2.73% +/- 0.85% vs. 0.89% +/- 0.26% retention from bolus, P = 0.04), which may underlie the lower hepatic lipid content. CONCLUSIONS: Here it is shown that HF diet negatively impacts postprandial TG clearance in offspring and results in an overall metabolic unfavorable phenotype and ectopic lipid deposition in the heart and in visceral storage sites.

AIMS/HYPOTHESIS: Human brown adipose tissue (BAT) has recently emerged as a potential target in the treatment of type 2 diabetes, owing to its capacity to actively clear glucose from the circulation-at least upon cold exposure. The effects of insulin resistance on the capacity of human BAT to take up glucose are unknown. Prolonged fasting is known to induce insulin resistance in peripheral tissues in order to spare glucose for the brain. METHODS: We studied the effect of fasting-induced insulin resistance on the capacity of BAT to take up glucose during cold exposure as well as on cold-stimulated thermogenesis. BAT glucose uptake was assessed by means of cold-stimulated dynamic 2-deoxy-2-[18F]fluoro-D-glucose positron emission tomography/computed tomography ([18F]FDG-PET/CT) imaging. RESULTS: We show that a 54 h fasting period markedly decreases both cold-induced BAT glucose uptake and nonshivering thermogenesis (NST) during cold stimulation. In vivo molecular imaging and modelling revealed that the reduction of glucose uptake in BAT was due to impaired cellular glucose uptake and not due to decreased supply. Interestingly, decreased BAT glucose uptake upon fasting was related to a decrease in core temperature during cold exposure, pointing towards a role for BAT in maintaining normothermia in humans. CONCLUSIONS/INTERPRETATION: Cold-stimulated glucose uptake in BAT is strongly reduced upon prolonged fasting. When cold-stimulated glucose uptake in BAT is also reduced under other insulin-resistant states, such as diabetes, cold-induced activation of BAT may not be a valid way to improve glucose clearance by BAT under such conditions. Trial registration: www.trialregister.nl NTR3523 Funding: This work was supported by the EU FP7 project DIABAT (HEALTH-F2-2011-278373 to WDvML) and by the Netherlands Organization for Scientific Research (TOP 91209037 to WDvML).

Item does not contain fulltext, OBJECTIVE: Parental high-fat feeding was proposed to negatively impact metabolic health in offspring. Here, the ectopic fat storage in heart and liver in offspring was investigated, and the effects on mitochondrial function, de novo lipogenesis, and postprandial lipid metabolism were explored in detail. METHODS: Male and female mice received either a high-fat (HF) or standard chow (LF) diet during mating, gestation and lactation. All offspring animals received the HF diet. RESULTS: Abdominal visceral adipose tissue tended to be higher in HF/HF mice. Cardiac lipid content was also higher in the HF/HF mice (LF/HF vs. HF/HF: 1.03% +/- 0.08% vs. 1.33% +/- 0.07% of water signal, P = 0.01). In contrast, hepatic lipid content tended to be lower in HF/HF mice compared to LF/HF mice. A severely disturbed postprandial lipid clearance was revealed in HF/HF mice by the results from the triglyceride (TG) tolerance tests (LF/HF vs. HF/HF: 6,753 +/- 2,213 vs. 14,367 +/- 1,978 mmol l(-1) min(-1) , P = 0.01) and (13) C-fatty acid retention test (LF/HF vs. HF/HF: 2.73% +/- 0.85% vs. 0.89% +/- 0.26% retention from bolus, P = 0.04), which may underlie the lower hepatic lipid content. CONCLUSIONS: Here it is shown that HF diet negatively impacts postprandial TG clearance in offspring and results in an overall metabolic unfavorable phenotype and ectopic lipid deposition in the heart and in visceral storage sites.

Item does not contain fulltext, OBJECTIVE: Parental high-fat feeding was proposed to negatively impact metabolic health in offspring. Here, the ectopic fat storage in heart and liver in offspring was investigated, and the effects on mitochondrial function, de novo lipogenesis, and postprandial lipid metabolism were explored in detail. METHODS: Male and female mice received either a high-fat (HF) or standard chow (LF) diet during mating, gestation and lactation. All offspring animals received the HF diet. RESULTS: Abdominal visceral adipose tissue tended to be higher in HF/HF mice. Cardiac lipid content was also higher in the HF/HF mice (LF/HF vs. HF/HF: 1.03% +/- 0.08% vs. 1.33% +/- 0.07% of water signal, P = 0.01). In contrast, hepatic lipid content tended to be lower in HF/HF mice compared to LF/HF mice. A severely disturbed postprandial lipid clearance was revealed in HF/HF mice by the results from the triglyceride (TG) tolerance tests (LF/HF vs. HF/HF: 6,753 +/- 2,213 vs. 14,367 +/- 1,978 mmol l(-1) min(-1) , P = 0.01) and (13) C-fatty acid retention test (LF/HF vs. HF/HF: 2.73% +/- 0.85% vs. 0.89% +/- 0.26% retention from bolus, P = 0.04), which may underlie the lower hepatic lipid content. CONCLUSIONS: Here it is shown that HF diet negatively impacts postprandial TG clearance in offspring and results in an overall metabolic unfavorable phenotype and ectopic lipid deposition in the heart and in visceral storage sites.

OBJECTIVE: Parental high-fat feeding was proposed to negatively impact metabolic health in offspring. Here, the ectopic fat storage in heart and liver in offspring was investigated, and the effects on mitochondrial function, de novo lipogenesis, and postprandial lipid metabolism were explored in detail. METHODS: Male and female mice received either a high-fat (HF) or standard chow (LF) diet during mating, gestation and lactation. All offspring animals received the HF diet. RESULTS: Abdominal visceral adipose tissue tended to be higher in HF/HF mice. Cardiac lipid content was also higher in the HF/HF mice (LF/HF vs. HF/HF: 1.03% +/- 0.08% vs. 1.33% +/- 0.07% of water signal, P = 0.01). In contrast, hepatic lipid content tended to be lower in HF/HF mice compared to LF/HF mice. A severely disturbed postprandial lipid clearance was revealed in HF/HF mice by the results from the triglyceride (TG) tolerance tests (LF/HF vs. HF/HF: 6,753 +/- 2,213 vs. 14,367 +/- 1,978 mmol l(-1) min(-1) , P = 0.01) and (13) C-fatty acid retention test (LF/HF vs. HF/HF: 2.73% +/- 0.85% vs. 0.89% +/- 0.26% retention from bolus, P = 0.04), which may underlie the lower hepatic lipid content. CONCLUSIONS: Here it is shown that HF diet negatively impacts postprandial TG clearance in offspring and results in an overall metabolic unfavorable phenotype and ectopic lipid deposition in the heart and in visceral storage sites.

AIMS/HYPOTHESIS: Human brown adipose tissue (BAT) has recently emerged as a potential target in the treatment of type 2 diabetes, owing to its capacity to actively clear glucose from the circulation-at least upon cold exposure. The effects of insulin resistance on the capacity of human BAT to take up glucose are unknown. Prolonged fasting is known to induce insulin resistance in peripheral tissues in order to spare glucose for the brain. METHODS: We studied the effect of fasting-induced insulin resistance on the capacity of BAT to take up glucose during cold exposure as well as on cold-stimulated thermogenesis. BAT glucose uptake was assessed by means of cold-stimulated dynamic 2-deoxy-2-[18F]fluoro-D-glucose positron emission tomography/computed tomography ([18F]FDG-PET/CT) imaging. RESULTS: We show that a 54 h fasting period markedly decreases both cold-induced BAT glucose uptake and nonshivering thermogenesis (NST) during cold stimulation. In vivo molecular imaging and modelling revealed that the reduction of glucose uptake in BAT was due to impaired cellular glucose uptake and not due to decreased supply. Interestingly, decreased BAT glucose uptake upon fasting was related to a decrease in core temperature during cold exposure, pointing towards a role for BAT in maintaining normothermia in humans. CONCLUSIONS/INTERPRETATION: Cold-stimulated glucose uptake in BAT is strongly reduced upon prolonged fasting. When cold-stimulated glucose uptake in BAT is also reduced under other insulin-resistant states, such as diabetes, cold-induced activation of BAT may not be a valid way to improve glucose clearance by BAT under such conditions. Trial registration: www.trialregister.nl NTR3523 Funding: This work was supported by the EU FP7 project DIABAT (HEALTH-F2-2011-278373 to WDvML) and by the Netherlands Organization for Scientific Research (TOP 91209037 to WDvML).

The accumulation of lipids in non-adipose tissues is attracting increasing attention due to its correlation with obesity. In muscle tissue, ectopic deposition of specific lipids is further correlated with pathogenic development of insulin resistance and type 2 diabetes. Most intramyocellular lipids are organized into lipid droplets (LDs), which are metabolically active organelles. In order to better understand the putative role of LDs in pathogenesis, insight into both the location of LDs and nearby chemistry of muscle tissue is very useful. Here, we demonstrate the use of label-free coherent anti-Stokes Raman scattering (CARS) microscopy in combination with multivariate, chemometric analysis to visualize intracellular lipid accumulations in ex vivo muscle tissue. Consistent with our previous results, hyperspectral CARS microscopy showed an increase in LDs in tissues where LD proteins were overexpressed, and further chemometric analysis showed additional features morphologically (and chemically) similar to mitochondria that colocalized with LDs. CARS imaging is shown to be a very useful method for label-free stratification of ectopic fat deposition and cellular organelles in fresh tissue sections with virtually no sample preparation.

The accumulation of lipids in non-adipose tissues is attracting increasing attention due to its correlation with obesity. In muscle tissue, ectopic deposition of specific lipids is further correlated with pathogenic development of insulin resistance and type 2 diabetes. Most intramyocellular lipids are organized into lipid droplets (LDs), which are metabolically active organelles. In order to better understand the putative role of LDs in pathogenesis, insight into both the location of LDs and nearby chemistry of muscle tissue is very useful. Here, we demonstrate the use of label-free coherent anti-Stokes Raman scattering (CARS) microscopy in combination with multivariate, chemometric analysis to visualize intracellular lipid accumulations in ex vivo muscle tissue. Consistent with our previous results, hyperspectral CARS microscopy showed an increase in LDs in tissues where LD proteins were overexpressed, and further chemometric analysis showed additional features morphologically (and chemically) similar to mitochondria that colocalized with LDs. CARS imaging is shown to be a very useful method for label-free stratification of ectopic fat deposition and cellular organelles in fresh tissue sections with virtually no sample preparation.

Contains fulltext : 117721.pdf (publisher's version ) (Closed access), Primary B-cell lymphoma of the testis, breast and thyroid are rare and data concerning cytogenetic aberrations at these extranodal sites are scarce. We examined the presence of extranodal marginal zone lymphoma-associated translocations, t(11;18)(q21;q21), t(1;14)(p22;q32), t(14;18)(q32;q21), t(3;14)(p14.1;q32) and numerical aberrations of chromosomes 1, 3, 12 and 18 by fluorescence in situ hybridization in 6 extranodal marginal zone lymphomas and 24 diffuse large B-cell lymphomas with (n=9) or without (n=15) marginal zone lymphoma components, with primary localizations in the breast (n=15), testis (n=9) and thyroid (n=6). We found t(14;18)(q32;q21), with breakpoints in IGH and MALT1, in one testicular diffuse large B-cell lymphoma and in two diffuse large B-cell lymphomas of the breast. No other translocations, amplifications or deletions involving IGH, BCL-10, BCL-2, MALT1 and IAP2 were detected. Numerical aberrations occurred in 67% of the lymphomas, 67% of extranodal marginal zone lymphomas, 56% of diffuse large B-cell lymphomas with marginal zone lymphoma components and in 73% of 'de novo' diffuse large B-cell lymphomas. These included 78% of testis, 67% of thyroid and 60% of breast lymphomas, and included mainly trisomy 18 (n=16), trisomy 3 (n=8) and trisomy 1 (n=3). One testicular diffuse large B-cell lymphoma harbored both t(14;18)(q32;q21) and trisomy 3. Our results indicate that at least a few cases of diffuse large B-cell lymphoma of the testis and the breast belong to the spectrum of extranodal marginal zone lymphoma.

Contains fulltext : 117721.pdf (publisher's version ) (Closed access), Primary B-cell lymphoma of the testis, breast and thyroid are rare and data concerning cytogenetic aberrations at these extranodal sites are scarce. We examined the presence of extranodal marginal zone lymphoma-associated translocations, t(11;18)(q21;q21), t(1;14)(p22;q32), t(14;18)(q32;q21), t(3;14)(p14.1;q32) and numerical aberrations of chromosomes 1, 3, 12 and 18 by fluorescence in situ hybridization in 6 extranodal marginal zone lymphomas and 24 diffuse large B-cell lymphomas with (n=9) or without (n=15) marginal zone lymphoma components, with primary localizations in the breast (n=15), testis (n=9) and thyroid (n=6). We found t(14;18)(q32;q21), with breakpoints in IGH and MALT1, in one testicular diffuse large B-cell lymphoma and in two diffuse large B-cell lymphomas of the breast. No other translocations, amplifications or deletions involving IGH, BCL-10, BCL-2, MALT1 and IAP2 were detected. Numerical aberrations occurred in 67% of the lymphomas, 67% of extranodal marginal zone lymphomas, 56% of diffuse large B-cell lymphomas with marginal zone lymphoma components and in 73% of 'de novo' diffuse large B-cell lymphomas. These included 78% of testis, 67% of thyroid and 60% of breast lymphomas, and included mainly trisomy 18 (n=16), trisomy 3 (n=8) and trisomy 1 (n=3). One testicular diffuse large B-cell lymphoma harbored both t(14;18)(q32;q21) and trisomy 3. Our results indicate that at least a few cases of diffuse large B-cell lymphoma of the testis and the breast belong to the spectrum of extranodal marginal zone lymphoma.

Contains fulltext : 117721.pdf (publisher's version ) (Closed access), Primary B-cell lymphoma of the testis, breast and thyroid are rare and data concerning cytogenetic aberrations at these extranodal sites are scarce. We examined the presence of extranodal marginal zone lymphoma-associated translocations, t(11;18)(q21;q21), t(1;14)(p22;q32), t(14;18)(q32;q21), t(3;14)(p14.1;q32) and numerical aberrations of chromosomes 1, 3, 12 and 18 by fluorescence in situ hybridization in 6 extranodal marginal zone lymphomas and 24 diffuse large B-cell lymphomas with (n=9) or without (n=15) marginal zone lymphoma components, with primary localizations in the breast (n=15), testis (n=9) and thyroid (n=6). We found t(14;18)(q32;q21), with breakpoints in IGH and MALT1, in one testicular diffuse large B-cell lymphoma and in two diffuse large B-cell lymphomas of the breast. No other translocations, amplifications or deletions involving IGH, BCL-10, BCL-2, MALT1 and IAP2 were detected. Numerical aberrations occurred in 67% of the lymphomas, 67% of extranodal marginal zone lymphomas, 56% of diffuse large B-cell lymphomas with marginal zone lymphoma components and in 73% of 'de novo' diffuse large B-cell lymphomas. These included 78% of testis, 67% of thyroid and 60% of breast lymphomas, and included mainly trisomy 18 (n=16), trisomy 3 (n=8) and trisomy 1 (n=3). One testicular diffuse large B-cell lymphoma harbored both t(14;18)(q32;q21) and trisomy 3. Our results indicate that at least a few cases of diffuse large B-cell lymphoma of the testis and the breast belong to the spectrum of extranodal marginal zone lymphoma.

We hypothesized that imagery training would improve the fast onset of neuromuscular activation and thereby fast knee extensor isometric torque development. Forty young healthy participants, not involved in strength training, were assigned to one of four groups: physical training, imagery training, placebo training or control. The three training groups had three 15 min sessions per week for 4 weeks, with a 90° knee angle but were tested also at 120°. At 90° knee angle, maximal torque increased (∼8%) similarly in all three training groups. The torque–time integral (contractile impulse) over the first 40 ms after torque onset (TTI40) increased (P < 0.05) after physical training (by ∼100%), but only at 90°. This increase was significantly different from the delta values (change pre to post) in the control and placebo groups, whereas delta values in the imagery group were similar to those in the placebo group. The increases in TTI40 following physical training were related (r 2 = 0.81, P < 0.05) to significant increases of knee extensor rectified surface EMG at torque onset (EMG40). In conclusion, only physical training led to a knee angle specific increase of contractile impulse that was significantly different from placebo and controls and that was related to improved onset of neuromuscular activation.

We hypothesized that imagery training would improve the fast onset of neuromuscular activation and thereby fast knee extensor isometric torque development. Forty young healthy participants, not involved in strength training, were assigned to one of four groups: physical training, imagery training, placebo training or control. The three training groups had three 15 min sessions per week for 4 weeks, with a 90° knee angle but were tested also at 120°. At 90° knee angle, maximal torque increased (∼8%) similarly in all three training groups. The torque–time integral (contractile impulse) over the first 40 ms after torque onset (TTI40) increased (P < 0.05) after physical training (by ∼100%), but only at 90°. This increase was significantly different from the delta values (change pre to post) in the control and placebo groups, whereas delta values in the imagery group were similar to those in the placebo group. The increases in TTI40 following physical training were related (r 2 = 0.81, P < 0.05) to significant increases of knee extensor rectified surface EMG at torque onset (EMG40). In conclusion, only physical training led to a knee angle specific increase of contractile impulse that was significantly different from placebo and controls and that was related to improved onset of neuromuscular activation.

Contains fulltext : 51698.pdf (publisher's version ) (Closed access), Global glomerulosclerosis can be divided in the vascular (obsolescent) type and the glomerulopathic (solidified) type. In biopsies from children with recurrent nephrotic syndrome owing to minimal change nephropathy (MCN), we noticed small, globally sclerosed glomeruli that appeared to be distinct from global glomerulosclerosis. These small sclerosed glomeruli are best described as involuted glomeruli. We have characterized these involuted glomeruli in detail. We studied biopsies of 18 children (11 male, 7 female) with frequently relapsing MCN and evaluated possible explanatory variables. The involuted glomeruli can be differentiated from the other types of global glomerulosclerosis. Most notable is the presence of vital podocytes and parietal epithelial cells, which have retained their staining characteristics, in between the matrix, and the absence of periglomerular and tubulo-interstitial fibrosis. We observed involuted glomeruli in 12 out of 18 biopsies; the median percentage of involuted glomeruli was 6% (range 0-33%). The percentage of involuted glomeruli correlated with age at renal biopsy and the interval between onset of disease and time of renal biopsy, but not with gender, age at onset of disease, or prednisone dose. Multivariate analysis revealed that the interval between onset of disease and time of renal biopsy was the only independent predictor. In conclusion, glomerular involution is a special form of global glomerulosclerosis. The absence of periglomerular and tubulo-interstitial fibrosis suggests a different pathogenesis. Glomerular involution is a slow process. The clinical data suggest that involution is related to the duration of the disease process.

Contains fulltext : 52870.pdf (publisher's version ) (Closed access), Enhanced proliferation of MDS progenitors is abrogated by increased apoptosis of their progeny in vivo. We investigated whether bone marrow mononuclear cells (BMMNC) of MDS patients also showed enhanced proliferation and apoptosis in vitro in comparison with acute myeloid leukemia (AML) and normal BM (NBM). NBM showed a decrease in the number of clusters in time due to apoptosis of clusters and due to development of clusters into colonies with low apoptotic level. In MDS patients, about two-fold more clusters have developed at day 4, and in contrast with NBM, the total number of clusters at day 7 remained high in spite of an increasing percentage of apoptotic clusters (from 52 to 76%) in combination with more colony formation. The number of clusters and colonies showed a sharp decrease at day 10 because of persistently high apoptosis at cluster level and increasing apoptosis in colonies. BMMNC of AML patients showed a decreased proliferation with enhanced apoptosis at cluster level in contrast to a relatively low apoptotic levels in the colony-forming cells. This data show that increased proliferation is abrogated by enhanced apoptosis in MDS, whereas AML showed decreased proliferation with a low level of apoptosis in colony-forming cells. These growth profiles of BMMNC are independent of stromal influences and may represent intrinsic features of the MDS progenitors and accessory cell interactions.

Contains fulltext : 51698.pdf (publisher's version ) (Closed access), Global glomerulosclerosis can be divided in the vascular (obsolescent) type and the glomerulopathic (solidified) type. In biopsies from children with recurrent nephrotic syndrome owing to minimal change nephropathy (MCN), we noticed small, globally sclerosed glomeruli that appeared to be distinct from global glomerulosclerosis. These small sclerosed glomeruli are best described as involuted glomeruli. We have characterized these involuted glomeruli in detail. We studied biopsies of 18 children (11 male, 7 female) with frequently relapsing MCN and evaluated possible explanatory variables. The involuted glomeruli can be differentiated from the other types of global glomerulosclerosis. Most notable is the presence of vital podocytes and parietal epithelial cells, which have retained their staining characteristics, in between the matrix, and the absence of periglomerular and tubulo-interstitial fibrosis. We observed involuted glomeruli in 12 out of 18 biopsies; the median percentage of involuted glomeruli was 6% (range 0-33%). The percentage of involuted glomeruli correlated with age at renal biopsy and the interval between onset of disease and time of renal biopsy, but not with gender, age at onset of disease, or prednisone dose. Multivariate analysis revealed that the interval between onset of disease and time of renal biopsy was the only independent predictor. In conclusion, glomerular involution is a special form of global glomerulosclerosis. The absence of periglomerular and tubulo-interstitial fibrosis suggests a different pathogenesis. Glomerular involution is a slow process. The clinical data suggest that involution is related to the duration of the disease process.

Contains fulltext : 52870.pdf (publisher's version ) (Closed access), Enhanced proliferation of MDS progenitors is abrogated by increased apoptosis of their progeny in vivo. We investigated whether bone marrow mononuclear cells (BMMNC) of MDS patients also showed enhanced proliferation and apoptosis in vitro in comparison with acute myeloid leukemia (AML) and normal BM (NBM). NBM showed a decrease in the number of clusters in time due to apoptosis of clusters and due to development of clusters into colonies with low apoptotic level. In MDS patients, about two-fold more clusters have developed at day 4, and in contrast with NBM, the total number of clusters at day 7 remained high in spite of an increasing percentage of apoptotic clusters (from 52 to 76%) in combination with more colony formation. The number of clusters and colonies showed a sharp decrease at day 10 because of persistently high apoptosis at cluster level and increasing apoptosis in colonies. BMMNC of AML patients showed a decreased proliferation with enhanced apoptosis at cluster level in contrast to a relatively low apoptotic levels in the colony-forming cells. This data show that increased proliferation is abrogated by enhanced apoptosis in MDS, whereas AML showed decreased proliferation with a low level of apoptosis in colony-forming cells. These growth profiles of BMMNC are independent of stromal influences and may represent intrinsic features of the MDS progenitors and accessory cell interactions.

Contains fulltext : 51698.pdf (publisher's version ) (Closed access), Global glomerulosclerosis can be divided in the vascular (obsolescent) type and the glomerulopathic (solidified) type. In biopsies from children with recurrent nephrotic syndrome owing to minimal change nephropathy (MCN), we noticed small, globally sclerosed glomeruli that appeared to be distinct from global glomerulosclerosis. These small sclerosed glomeruli are best described as involuted glomeruli. We have characterized these involuted glomeruli in detail. We studied biopsies of 18 children (11 male, 7 female) with frequently relapsing MCN and evaluated possible explanatory variables. The involuted glomeruli can be differentiated from the other types of global glomerulosclerosis. Most notable is the presence of vital podocytes and parietal epithelial cells, which have retained their staining characteristics, in between the matrix, and the absence of periglomerular and tubulo-interstitial fibrosis. We observed involuted glomeruli in 12 out of 18 biopsies; the median percentage of involuted glomeruli was 6% (range 0-33%). The percentage of involuted glomeruli correlated with age at renal biopsy and the interval between onset of disease and time of renal biopsy, but not with gender, age at onset of disease, or prednisone dose. Multivariate analysis revealed that the interval between onset of disease and time of renal biopsy was the only independent predictor. In conclusion, glomerular involution is a special form of global glomerulosclerosis. The absence of periglomerular and tubulo-interstitial fibrosis suggests a different pathogenesis. Glomerular involution is a slow process. The clinical data suggest that involution is related to the duration of the disease process.

Contains fulltext : 57907.pdf (publisher's version ) (Closed access), OBJECTIVE: To investigate whether the detection of proliferation-associated Ki-67 antigen may be of value in differentiating between reserve cell hyperplasia (RCH) and small cell lung cancer (SCLC). STUDY DESIGN: Retrospectively, 20 Papanicolaou-stained bronchial brushes or washings from 20 patients were selected. Ten were diagnosed as RCH (and had no SCLC in follow-up) and the other 10 as SCLC (histologically confirmed). All 20 Papanicolaou-stained slides were restained with the monoclonal antibody MIB1, directed against Ki-67 antigen; that simple and reliable procedure was described recently. In each specimen 5 coherent cell groups were identified, corresponding to RCH or SCLC, respectively; photographed; and studied for Ki-67 antigen expression after MIB1 staining of the slides. At least 3 cell groups remained in each specimen. The Ki-67 labeling index (LI) of the specimens was determined as the number of MIB1-positive cells divided by the total number of cells in the remaining cell groups. RESULTS: All cases of SCLC showed a mean Ki-67 LI of at least .415 (mean .684, SD .151), whereas in the cases with RCH the mean Ki-67 LI never was more than .158 (mean .048, SD .049). The difference was highly significant (P<.001, Student's t test). Linear discriminant analysis resulted in a classifier with which we were able to discriminate correctly between SCLC and RCH in 100% of the 20 bronchial brushings and washings. CONCLUSION: The results clearly demonstrate that measuring proliferative activity in Papanicolaou-stained bronchial brushings and washings by MIB1 restaining of the slides may be of great practical value in accurately discriminating RCH from SCLC. The method is simple and can be performed in any laboratory that is able to carry out immunocytochemical staining. However, an additional (prospective) study with a series of difficult cases is necessary to confirm these findings.

Contains fulltext : 57907.pdf (publisher's version ) (Closed access), OBJECTIVE: To investigate whether the detection of proliferation-associated Ki-67 antigen may be of value in differentiating between reserve cell hyperplasia (RCH) and small cell lung cancer (SCLC). STUDY DESIGN: Retrospectively, 20 Papanicolaou-stained bronchial brushes or washings from 20 patients were selected. Ten were diagnosed as RCH (and had no SCLC in follow-up) and the other 10 as SCLC (histologically confirmed). All 20 Papanicolaou-stained slides were restained with the monoclonal antibody MIB1, directed against Ki-67 antigen; that simple and reliable procedure was described recently. In each specimen 5 coherent cell groups were identified, corresponding to RCH or SCLC, respectively; photographed; and studied for Ki-67 antigen expression after MIB1 staining of the slides. At least 3 cell groups remained in each specimen. The Ki-67 labeling index (LI) of the specimens was determined as the number of MIB1-positive cells divided by the total number of cells in the remaining cell groups. RESULTS: All cases of SCLC showed a mean Ki-67 LI of at least .415 (mean .684, SD .151), whereas in the cases with RCH the mean Ki-67 LI never was more than .158 (mean .048, SD .049). The difference was highly significant (P<.001, Student's t test). Linear discriminant analysis resulted in a classifier with which we were able to discriminate correctly between SCLC and RCH in 100% of the 20 bronchial brushings and washings. CONCLUSION: The results clearly demonstrate that measuring proliferative activity in Papanicolaou-stained bronchial brushings and washings by MIB1 restaining of the slides may be of great practical value in accurately discriminating RCH from SCLC. The method is simple and can be performed in any laboratory that is able to carry out immunocytochemical staining. However, an additional (prospective) study with a series of difficult cases is necessary to confirm these findings.

Contains fulltext : 57907.pdf (publisher's version ) (Closed access), OBJECTIVE: To investigate whether the detection of proliferation-associated Ki-67 antigen may be of value in differentiating between reserve cell hyperplasia (RCH) and small cell lung cancer (SCLC). STUDY DESIGN: Retrospectively, 20 Papanicolaou-stained bronchial brushes or washings from 20 patients were selected. Ten were diagnosed as RCH (and had no SCLC in follow-up) and the other 10 as SCLC (histologically confirmed). All 20 Papanicolaou-stained slides were restained with the monoclonal antibody MIB1, directed against Ki-67 antigen; that simple and reliable procedure was described recently. In each specimen 5 coherent cell groups were identified, corresponding to RCH or SCLC, respectively; photographed; and studied for Ki-67 antigen expression after MIB1 staining of the slides. At least 3 cell groups remained in each specimen. The Ki-67 labeling index (LI) of the specimens was determined as the number of MIB1-positive cells divided by the total number of cells in the remaining cell groups. RESULTS: All cases of SCLC showed a mean Ki-67 LI of at least .415 (mean .684, SD .151), whereas in the cases with RCH the mean Ki-67 LI never was more than .158 (mean .048, SD .049). The difference was highly significant (P<.001, Student's t test). Linear discriminant analysis resulted in a classifier with which we were able to discriminate correctly between SCLC and RCH in 100% of the 20 bronchial brushings and washings. CONCLUSION: The results clearly demonstrate that measuring proliferative activity in Papanicolaou-stained bronchial brushings and washings by MIB1 restaining of the slides may be of great practical value in accurately discriminating RCH from SCLC. The method is simple and can be performed in any laboratory that is able to carry out immunocytochemical staining. However, an additional (prospective) study with a series of difficult cases is necessary to confirm these findings.

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Contains fulltext : 24642___.PDF (publisher's version ) (Open Access)

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Contains fulltext : 24642___.PDF (publisher's version ) (Open Access)