To develop a color code and to investigate the validity of Laser Speckle Contrast Imaging (LSCI) for measuring burn wound healing potential (HP) in burn patients as compared to the reference standard ...Laser Doppler Imaging (LDI).
A prospective, observational, cohort study was conducted in adult patients with acute burn wounds. The relationship between mean flux measured with LDI and mean perfusion units (PU) measured with LSCI was expressed in a regression formula. Measurements were performed between 2 and 5 days after the burn wound. The creation of a LSCI color code was done by mapping the clinically validated color code of the LDI to the corresponding values on the LSCI scale. To assess validity of the LSCI, the ability of the LSCI to discriminate between HP < 14 and ≥ 14 days and HP < 21 and original ≥ 21 days according to the LDI reference standard was evaluated, with calculation of receiver operating characteristics (ROC) curves.
A total of 50 patients were included with a median age of 40 years and total body surface area burned of 6%. LSCI values of 143 PU and 113 PU were derived as the cut-off values for the need of conservative treatment (HP < 14 and ≥ 14 days) resp. surgical closure (HP < 21 and ≥ 21 days). These LSCI cut off values showed a good discrimination between HP 14 days versus ≥ 14 days (Area Under Curve (AUC)= 0.89; sensitivity 85% and specificity = 82%) and a good discrimination between HP 21 days versus ≥ 21 days (AUC of 0.89, sensitivity 81% and specificity 88%).
This is the first study in which a color code for the LSCI in adult clinical burn patients has been developed. Our study reconfirms the good performance of the LSCI for prediction of burn wound healing potential. This provides additional evidence for the potential value of the LSCI in specialized burn care.
Most patients with Post COVID Syndrome (PCS) present with a plethora of symptoms without clear evidence of organ dysfunction. A subset of them fulfills diagnostic criteria of myalgic ...encephalomyelitis/chronic fatigue syndrome (ME/CFS). Symptom severity of ME/CFS correlates with natural regulatory autoantibody (AAB) levels targeting several G-protein coupled receptors (GPCR). In this exploratory study, we analyzed serum AAB levels against vaso- and immunoregulatory receptors, mostly GPCRs, in 80 PCS patients following mild-to-moderate COVID-19, with 40 of them fulfilling diagnostic criteria of ME/CFS. Healthy seronegative (n=38) and asymptomatic post COVID-19 controls (n=40) were also included in the study as control groups. We found lower levels for various AABs in PCS compared to at least one control group, accompanied by alterations in the correlations among AABs. Classification using random forest indicated AABs targeting ADRB2, STAB1, and ADRA2A as the strongest classifiers (AABs stratifying patients according to disease outcomes) of post COVID-19 outcomes. Several AABs correlated with symptom severity in PCS groups. Remarkably, severity of fatigue and vasomotor symptoms were associated with ADRB2 AAB levels in PCS/ME/CFS patients. Our study identified dysregulation of AAB against various receptors involved in the autonomous nervous system (ANS), vaso-, and immunoregulation and their correlation with symptom severity, pointing to their role in the pathogenesis of PCS.
Peritoneal metastases (PM) in colorectal cancer (CRC) are associated with therapy resistance and poor survival. Oxaliplatin monotherapy is widely applied in the intraperitoneal treatment of PM, but ...fails to yield clinical benefit. We aimed to identify the mechanism(s) underlying PM resistance to oxaliplatin and to develop strategies overcoming such resistance.
We generated a biobank consisting of 35 primary tumour regions and 59 paired PM from 12 patients. All samples were analysed by RNA sequencing. We also generated a series of PM-derived organoid (PMDO) cultures and used these to design and test strategies to overcome resistance to oxaliplatin.
PM displayed various hallmarks of aggressive CRC biology. The vast majority of PM and paired primary tumours belonged to the Consensus Molecular Subtype 4 (CMS4). PMDO cultures were resistant to oxaliplatin and expressed high levels of glutamate-cysteine ligase (GCLC) causing detoxification of oxaliplatin through glutathione synthesis. Genetic or pharmacological targeting of GCLC sensitised PMDOs to a 1-h exposure to oxaliplatin, through increased platinum-DNA adduct formation.
These results link oxaliplatin resistance of colorectal PM to their CMS4 status and high reducing capacity. Inhibiting the reducing capacity of PM may be an effective strategy to overcome PM resistance to oxaliplatin.
A high-fat diet has been recommended for the treatment of patients with mitochondrial myopathy due to complex I (NADH dehydrogenase) deficiency (CID).
This study evaluated the effects of intravenous ...infusion of isoenergetic amounts of triacylglycerol or glucose on substrate oxidation, glycolytic carbohydrate metabolism, and exercise endurance time and energy state of muscle in CID patients.
Four CID patients and 15 control subjects were infused with triacylglycerol (3.7 mg x kg(-1) x min(-1)) or glucose (10 mg x kg(-1) x min(-1)) during low-intensity leg exercise. Respiratory calorimetry was used to evaluate mitochondrial substrate oxidation. The concentration and rate of appearance of plasma lactate (from dilution of 1-(13)Clactate) were used to evaluate glycolytic carbohydrate metabolism. (31)P magnetic resonance spectroscopy was used to determine ratios of phosphocreatine to inorganic o-phosphate in forearm muscle during exercise.
In 3 patients, leg exercise endurance time was better during the triacylglycerol infusion than during the glucose infusion. In all 4 patients, whole-body oxygen consumption rates during exercise were higher during triacylglycerol infusion than during the glucose infusion. In 3 patients, the concentration and rate of appearance of plasma lactate were lower during triacylglycerol infusion than during the glucose infusion. Ratios of phosphocreatine to inorganic o-phosphate during exercise were not significantly different between the 2 infusion studies or between the patients and control subjects.
Triacylglycerol infusion is associated with a greater oxidation of substrates, lower rates of appearance and concentrations of plasma lactate, and greater leg exercise endurance time in myopathic CID patients than is glucose infusion. The energy state of muscle during exercise, however, was not significantly different after infusion of triacylglycerol or glucose.
The urinary loss of transferrin is sufficient to reduce plasma transferrin concentrations in the nephrotic syndrome. Hypotransferrinemia may lead to iron loss and microcytic anemia. The mechanism ...responsible for the hypotransferrinemia in the nephrotic syndrome is, however, unknown. In the present study, synthesis rate of transferrin was measured in vivo in nephrotic patients (n = 7) compared with control subjects (n = 6) using L-1-(13)C-valine. Plasma transferrin and iron concentration in the patients were significantly lower than in control subjects (transferrin, 1.39 +/- 0.08 versus 2.57 +/- 0.11 g/L, P < 0.0001; iron, 10.2 +/- 0.8 versus 21.1 +/- 4.5 micromol/L, P = 0.02). Furthermore, albuminuria correlated with transferrinuria (r(2) = 0.901, P = 0.001). The absolute synthesis rate of transferrin was increased in the patients (10.0 +/- 1.1 versus 7.4 +/- 0.7 mg/kg per d, P = 0.07), although this value failed to achieve significance. C-reactive protein, plasma iron, and proteinuria did not correlate with transferrin synthesis. In contrast, transferrin synthesis correlated with albumin synthesis (r(2) = 0.648, P = 0.03; n = 7). The present study indicates that increased transferrin synthesis occurs in nephrotic patients but is insufficient to compensate for urinary losses. Because, overall, no significant relationship was found between transferrin synthesis and either C-reactive protein or iron, it is unlikely that inflammation suppresses or that iron deficiency stimulates increased transferrin synthesis in these patients. The correlation between transferrin synthesis and albumin synthesis suggests that transferrin synthesis is a component of a general response in hepatic protein synthesis in the nephrotic syndrome. This suggests that a therapeutic approach to maximize plasma transferrin concentrations in nephrotic patients should be aimed primarily at reducing urinary protein excretion.
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.
LDs were stained by MDH together with combinations of mitochondrial 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.
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.
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.
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•MDH can be used as a blue lipid droplet dye in tissue sections.•PLIN5 is located in the cytosol and at the lipid droplet mitochondrial interface.•Super-resolution microscopy reveals that PLIN5 is not at the mitochondrial cristae.•PLIN2 and PLIN5 localize to the lipid droplet at distinct locations rather than fully decorating the lipid droplet.
Aims/hypothesis
In contrast to insulin-resistant individuals, insulin-sensitive athletes possess high intramyocellular lipid content (IMCL), good mitochondrial function and high perilipin 5 (PLIN5) ...levels, suggesting a role for PLIN5 in benign IMCL storage. We hypothesised a role for PLIN5 in modulating fasting-mediated insulin resistance.
Methods
Twelve men were fasted for 60 h, before and after which muscle biopsies were taken and stained for lipid droplets (LDs), PLIN5 and laminin. Confocal microscopy images were analysed for LD size, number, PLIN5 association and subcellular distribution.
Results
Fasting elevated IMCL content 2.8-fold and reduced insulin sensitivity (by 55%). Individuals with the most prominent increase in IMCL showed the least reduction in insulin sensitivity (
r
= 0.657;
p
= 0.028) and mitochondrial function (
r
= 0.896;
p
= 0.006). During fasting,
PLIN5
gene expression or PLIN5 protein content in muscle homogenates was unaffected, microscopy analyses revealed that the fraction of PLIN5 associated with LDs (PLIN5+) increased significantly (+26%) upon fasting, suggesting PLIN5 redistribution. The significant increase in LD number (+23%) and size (+23%) upon fasting was entirely accounted for by PLIN5+ LDs, not by LDs devoid of PLIN5. Also the association between IMCL storage capacity and insulin resistance and mitochondrial dysfunction was only apparent for PLIN5+ LDs.
Conclusions/interpretation
Fasting results in subcellular redistribution of PLIN5 and promotes the capacity to store excess fat in larger and more numerous PLIN5-decorated LDs. This associates with blunting of fasting-induced insulin resistance and mitochondrial dysfunction, suggesting a role for PLIN5 in the modulation of fasting-mediated lipotoxicity.
Trial registration
:
trialregister.nl NTR 2042
A multidisciplinary cognitive behavioral program might have beneficial effects on people with chronic neuropathic spinal cord injury pain.
Many people with spinal cord injury (SCI) rate chronic ...neuropathic pain as one of the most difficult problems to manage. The aim of the CONECSI (COping with NEuropathiC Spinal cord Injury pain) trial was to evaluate a multidisciplinary cognitive behavioral treatment program for persons with chronic neuropathic pain after SCI. The intervention consisted of educational, cognitive, and behavioral elements. A total of 61 people were randomized to either the intervention group or the waiting list control group in 4 Dutch rehabilitation centers. Primary outcomes were pain intensity and pain-related disability (Chronic Pain Grade questionnaire), and secondary outcomes were mood (Hospital Anxiety and Depression Scale), participation in activities (Utrecht Activities List), and life satisfaction (Life Satisfaction Questionnaire). Measurements were performed at baseline, and at 3, and 6
months follow-up. The primary statistical technique was random coefficient analysis. The analyses showed significant changes over time on both primary (t1–t2), and 2 out of 4 secondary outcomes (both t1–t2 and t1–t3). Significant intervention effects (Time
*
Group interactions) were found for anxiety and participation in activities, but not for the primary outcomes. Subsequent paired
t tests showed significant changes in the intervention group that were not seen in the control group: decrease of pain intensity, pain-related disability, anxiety, and increase of participation in activities. This study implies that a multidisciplinary cognitive behavioral program might have beneficial effects on people with chronic neuropathic SCI pain.
Laminopathies comprise a group of inherited diseases with variable clinical phenotypes, caused by mutations in the lamin A/C gene (LMNA). A prominent feature in several of these diseases is muscle ...wasting, as seen in Emery–Dreifuss muscle dystrophy, dilated cardiomyopathy and limb-girdle muscular dystrophy. Although the mechanisms underlying this phenotype remain largely obscure, two major working hypotheses are currently being investigated, namely, defects in gene regulation and/or abnormalities in nuclear architecture causing cellular fragility. In this study, using a newly developed cell compression device we have tested the latter hypothesis. The device allows controlled application of mechanical load onto single living cells, with simultaneous visualization of cellular deformation and quantitation of resistance. With the device, we have compared wild-type (MEF+/+) and LMNA knockout (MEF−/−) mouse embryonic fibroblasts (MEFs), and found that MEF−/− cells show a significantly decreased mechanical stiffness and a significantly lower bursting force. Partial rescue of the phenotype by transfection with either lamin A or lamin C prevented gross nuclear disruption, as seen in MEF−/− cells, but was unable to fully restore mechanical stiffness in these cells. Our studies show a direct correlation between absence of LMNA proteins and nuclear fragility in living cells. Simultaneous recordings by confocal microscopy revealed that the nuclei in MEF−/− cells, in contrast to MEF+/+ cells, exhibited an isotropic deformation upon indentation, despite an anisotropic deformation of the cell as a whole. This nuclear behaviour is indicative for a loss of interaction of the disturbed nucleus with the surrounding cytoskeleton. In addition, careful investigation of the three-dimensional organization of actin-, vimentin- and tubulin-based filaments showed a disturbed interaction of these structures in MEF−/− cells. Therefore, we suggest that in addition to the loss of nuclear stiffness, the loss of a physical interaction between nuclear structures (i.e. lamins) and the cytoskeleton is causing more general cellular weakness and emphasizes a potential key function for lamins in maintaining cellular tensegrity.
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