Platelet lysate as a serum replacement for skin cell culture on biomimetic PCL nanofibers

Platelets are a popular source of native growth factors for tissue engineering applications. The aim of the study was to verify the use of platelet lysate as a fetal bovine serum (FBS) replacement for skin cell culture. The cytokine content of the platelet lysate was characterized using the Bio-Plex system. The cells (fibroblasts, melanocytes, and keratinocytes) were cultured on PCL nanofibrous scaffolds to mimic their natural microenvironment. The cytokine content of the platelet lysate was determined, and to the cells, a medium containing platelet lysate or platelet lysate in combination with FBS was added. The results showed that 7% (v/v) platelet lysate was sufficient to supplement 10% (v/v) FBS in the culture of fibroblasts and keratinocytes. The combination of platelet lysate and FBS had a rather inhibitory effect on fibroblasts, in contrary to keratinocytes, where the effect was synergic. Platelet lysate did not sufficiently promote proliferation in melanocytes; however, the combination of FBS and platelet lysate yielded a better outcome and resulted in bipolar morphology of the cultured melanocytes. The data indicated that platelet lysate improved cell proliferation and metabolic activity and may be used as an additive to the cell culture media.     

A Novel Public MR Image Dataset of Multiple Sclerosis Patients With Lesion Segmentations Based on Multi-rater Consensus

Quantified volume and count of white-matter lesions based on magnetic resonance (MR) images are important biomarkers in several neurodegenerative diseases. For a routine extraction of these biomarkers an accurate and reliable automated lesion segmentation is required. To objectively and reliably determine a standard automated method, however, creation of standard validation datasets is of extremely high importance. Ideally, these datasets should be publicly available in conjunction with standardized evaluation methodology to enable objective validation of novel and existing methods. For validation purposes, we present a novel MR dataset of 30 multiple sclerosis patients and a novel protocol for creating reference white-matter lesion segmentations based on multi-rater consensus. On these datasets three expert raters individually segmented white-matter lesions, using in-house developed semi-automated lesion contouring tools. Later, the raters revised the segmentations in several joint sessions to reach a consensus on segmentation of lesions. To evaluate the variability, and as quality assurance, the protocol was executed twice on the same MR images, with a six months break. The obtained intra-consensus variability was substantially lower compared to the intra- and inter-rater variabilities, showing improved reliability of lesion segmentation by the proposed protocol. Hence, the obtained reference segmentations may represent a more precise target to evaluate, compare against and also train, the automatic segmentations. To encourage further use and research we will publicly disseminate on our website http://lit.fe.uni-lj.si/tools the tools used to create lesion segmentations, the original and preprocessed MR image datasets and the consensus lesion segmentations.     

News in the classification of pulmonary adenocarcinomas and potential prognostic and predictive factors in non-small lung cancer

Lung cancers are still divided into two major subgroups: small-cell and non-small cell lung cancer (NSCLC) irrespective of biological heterogeneity of NSCLC. It is a key task of the pathologist to provide an accurate classification of tumorous lesions to avoid the term NSCLC and to use it only in the vast minority of cases. Moreover, the most recent reclassification of pulmonary adenocarcinomas should be reflected in the standard biopsy protocol reporting. There is also an increasingly urgent need to provide high quality material for testing of the genetic characteristics of NSCLC, especially the presence and functional status of the EGFR receptor (epidermal growth factor receptor), as well as other potential prognostic markers. The requirement for the quality and swiftness of diagnosis puts major emphasis on the close multidisciplinary collaboration with the central role of a specialized pathologist, who coordinates the differential-diagnostic procedure. This in turn implies the necessity of accounting for the increasing financial burden of diagnostic departments.     

Impaired cardiac ischemic tolerance in spontaneously hypertensive rats is attenuated by adaptation to chronic and acute stress

Chronic hypertension may have a negative impact on the myocardial response to ischemia. On the other hand, intrinsic ischemic tolerance may persist even in the pathologically altered hearts of hypertensive animals, and may be modified by short- or long-term adaptation to different stressful conditions. The effects of long-term limitation of living space (ie, crowding stress [CS]) and brief ischemia-induced stress on cardiac response to ischemia/reperfusion (I/R) injury are not yet fully characterized in hypertensive subjects. The present study was designed to test the influence of chronic and acute stress on the myocardial response to I/R in spontaneously hypertensive rats (SHR) compared with their effects in normotensive counterparts. In both groups, chronic, eight-week CS was induced by caging five rats per cage in cages designed for two rats (200 cm²/rat), while controls (C) were housed four to a cage in cages designed for six animals (480 cm²/rat). Acute stress was evoked by one cycle of I/R (5 min each, ischemic preconditioning) before sustained I/R in isolated Langendorff-perfused hearts of normotensive and SHR rats. At baseline conditions, the effects of CS were manifested only as a further increase in blood pressure in SHR, and by marked limitation of coronary perfusion in normotensive animals, while no changes in heart mechanical function were observed in any of the groups. Postischemic recovery of contractile function, severity of ventricular arrhythmias and lethal injury (infarction size) were worsened in the hypertrophied hearts of C-SHR compared with normotensive C. However, myo-cardial stunning and reperfusion-induced ventricular arrhythmias were attenuated by CS in SHR, which was different from deterioration of I/R injury in the hearts of normotensive animals. In contrast, ischemic preconditioning conferred an effective protection against I/R in both groups, although the extent of anti-infarct and anti-arrhythmic effects was lower in SHR. Both forms of stress may improve the altered response to ischemia in hypertensive subjects. In contrast to short-term preconditioning stress, chronic psychosocial stress was associated with a higher risk of lethal arrhythmias and contractile failure in normotensive animals exposed to an acute ischemic challenge.     

Erythropoietin facilitates the return of spontaneous circulation and survival in victims of out-of-hospital cardiac arrest

Background

Erythropoietin activates potent protective mechanisms in non-hematopoietic tissues including the myocardium. In a rat model of ventricular fibrillation, erythropoietin preserved myocardial compliance enabling hemodynamically more effective CPR.

Objective

To investigate whether intravenous erythropoietin given within 2 min of physician-led CPR improves outcome from out-of-hospital cardiac arrest.

Methods

Erythropoietin (90,000 IU of beta-epoetin, n = 24) was compared prospectively with 0.9% NaCl (concurrent controls = 30) and retrospectively with a preceding group treated with similar protocol (matched controls = 48).

Results

Compared with concurrent controls, the erythropoietin group had higher rates of ICU admission (92% vs 50%, p = 0.004), return of spontaneous circulation (ROSC) (92% vs 53%, p = 0.006), 24-h survival (83% vs 47%, p = 0.008), and hospital survival (54% vs 20%, p = 0.011). However, after adjusting for pretreatment covariates only ICU admission and ROSC remained statistically significant. Compared with matched controls, the erythropoietin group had higher rates of ICU admission (92% vs 65%, p = 0.024) and 24-h survival (83% vs 52%, p = 0.014) with statistically insignificant higher ROSC (92% vs 71%, p = 0.060) and hospital survival (54% vs 31%, p = 0.063). However, after adjusting for pretreatment covariates all four outcomes were statistically significant. End-tidal PCO₂ (an estimate of blood flow during chest compression) was higher in the erythropoietin group.

Conclusions

Erythropoietin given during CPR facilitates ROSC, ICU admission, 24-h survival, and hospital survival. This effect was consistent with myocardial protection leading to hemodynamically more effective CPR (Trial registration: http://isrctn.org. Identifier: ISRCTN67856342).     

Near-infrared spectroscopy during stagnant ischemia estimates central venous oxygen saturation and mixed venous oxygen saturation discrepancy in patients with severe left heart failure and additional sepsis/septic shock

Introduction

Discrepancies of 5-24% between superior vena cava oxygen saturation (ScvO₂) and mixed venous oxygen saturation (SvO₂) have been reported in patients with severe heart failure. Thenar muscle tissue oxygenation (StO₂) measured with near-infrared spectroscopy (NIRS) during arterial occlusion testing decreases slower in sepsis/septic shock patients (lower StO₂ deoxygenation rate). The StO₂ deoxygenation rate is influenced by dobutamine. The aim of this study was to determine the relationship between the StO₂ deoxygenation rate and the ScvO₂-SvO₂ discrepancy in patients with severe left heart failure and additional sepsis/septic shock treated with or without dobutamine.

Methods

Fifty-two patients with severe left heart failure due to primary heart disease with additional severe sepsis/septic shock were included. SvO₂ and ScvO₂ were compared to the thenar muscle StO₂ before and during arterial occlusion.

Results

SvO₂ correlated significantly with ScvO₂ (Pearson correlation 0.659, P = 0.001), however, Bland Altman analysis showed a clinically important difference between both variables (ScvO₂-SvO₂ mean 72 ± 8%, ScvO₂-SvO₂ difference 9.4 ± 7.5%). The ScvO₂-SvO₂ difference correlated with plasma lactate (Pearson correlation 0.400, P = 0.003) and the StO₂ deoxygenation rate (Pearson correlation 0.651, P = 0.001). In the group of patients treated with dobutamine, the ScvO₂-SvO₂ difference correlated with plasma lactate (Pearson correlation 0.389, P = 0.011) and the StO₂ deoxygenation rate (Pearson correlation 0.777, P = 0.0001).

Conclusions

In patients with severe heart failure with additional severe sepsis/septic shock the ScvO₂-SvO₂ discrepancy presents a clinical problem. In these patients the skeletal muscle StO₂ deoxygenation rate is inversely proportional to the difference between ScvO₂ and SvO₂; dobutamine does not influence this relationship. When using ScvO₂ as a treatment goal, the NIRS measurement may prove to be a useful non-invasive diagnostic test to uncover patients with a normal ScvO₂ but potentially an abnormally low SvO₂.

Trial Registration

{"type":"clinical-trial","attrs":{"text":"NCT00384644","term_id":"NCT00384644"}}NCT00384644 ClinicalTrials.Gov.     

Monounsaturated fatty acids in serum triacylglycerols are associated with response to neoadjuvant chemotherapy in breast cancer patients

Changes in cellular lipid metabolism are a common feature in most solid tumors, which occur already in early stages of the tumor progression. However, it remains unclear if the tumor‐specific lipid changes can be detected at the level of systemic lipid metabolism. The objective of this study was to perform comprehensive analysis of lipids in breast cancer patient serum samples. Lipidomic profiling using an established analytical platform was performed in two cohorts of breast cancer patients receiving neoadjuvant chemotherapy. The analyses were performed for 142 patients before and after neoadjuvant chemotherapy, and the results before chemotherapy were validated in an independent cohort of 194 patients. The analyses revealed that in general the tumor characteristics are not reflected in the serum samples. However, there was an association of specific triacylglycerols (TGs) in patients' response to chemotherapy. These TGs containing mainly oleic acid (C18:1) were found in lower levels in those patients showing pathologic complete response before receiving chemotherapy. Some of these TGs were also associated with estrogen receptor status and overall or disease‐free survival of the patients. The results suggest that the altered serum levels of oleic acid in breast cancer patients are associated with their response to chemotherapy.     

Inhibition of sex hormone-binding globulin production in the human hepatoma (Hep G2) cell line by insulin and prolactin

Sex hormone-binding globulin (SHBG) production in humans has been thought to be stimulated by estrogens and thyroid hormone and inhibited by androgens. However, recent data indicate that SHBG production in vitro is stimulated by both androgens and estrogens. This study was designed to determine what other hormonal factors regulate SHBG production. Since hyperinsulinemia and hyperprolactinemia both occur in disease states in which low serum SHBG levels are found, the effects of insulin and PRL were compared to and/or studied in combination with estradiol (E2), T4, and testosterone (T) in a human hepatoma cell line (Hep G2). Hep G2 cells were grown to near confluence in medium including 10% fetal calf serum, and then 72-h experimental incubations were carried out which used only fetal calf serum-free medium. Compared to control incubations, both insulin (10(-8) mol/L) and PRL (10(-8) mol/L) decreased SHBG production from 65.0 +/- 0.6 (+/- SE) to 46.8 +/- 1.1 and 46.8 +/- 1.2 nmol/10(6) cells, respectively (P less than 0.01). Insulin also inhibited both E2 and T4-stimulated SHBG production. T stimulated SHBG production to the same degree as E2. Finally, both E2 and insulin significantly increased cell number, an important consideration when expressing the effect of a hormone on SHBG production in cultured cells. We conclude that insulin and PRL inhibit SHBG production and confirm that T4, T, and E2 stimulate SHBG production in vitro. These findings suggest that insulin and PRL may be important factors in the regulation of SHBG production in vivo.