Development, characterization and sterilisation of Nanocellulose-alginate-(hyaluronic acid)- bioinks and 3D bioprinted scaffolds for tissue engineering.

3D-bioprinting is an emerging technology of high potential in tissue engineering (TE), since it shows effective control over scaffold fabrication and cell distribution. Biopolymers such as alginate (Alg), nanofibrillated cellulose (NC) and hyaluronic acid (HA) offer excellent characteristics for use as bioinks due to their excellent biocompatibility and rheological properties. Cell incorporation into the bioink requires sterilisation assurance, and autoclave, ß-radiation and γ-radiation are widely used sterilisation techniques in biomedicine; however, their use in 3D-bioprinting for bioinks sterilisation is still in their early stages. In this study, different sterilisation procedures were applied on NC-Alg and NC-Alg-HA bioinks and their effect on several parameters was evaluated. Results demonstrated that NC-Alg and NC-Alg-HA bioinks suffered relevant rheological and physicochemical modifications after sterilisation; yet, it can be concluded that the short cycle autoclave is the best option to sterilise both NC-Alg based cell-free bioinks, and that the incorporation of HA to the NC-Alg bioink improves its characteristics. Additionally, 3D scaffolds were bioprinted and specifically characterized as well as the D1 mesenchymal stromal cells (D1-MSCs) embedded for cell viability analysis. Notably, the addition of HA demonstrates better scaffold properties, together with higher biocompatibility and cell viability in comparison with the NC-Alg scaffolds. Thus, the use of MSCs containing NC-Alg based scaffolds may become a feasible tissue engineering approach for regenerative medicine.

Variability and effects of bronchial colonisation in patients with moderate COPD.

Sputum and lung function were periodically assessed in stable moderate chronic obstructive pulmonary disease (COPD) outpatients to determine relationships between bronchial colonisation and inflammation. Relationships between potentially pathogenic microorganism (PPM) typology, bronchial inflammation (neutrophilia, tumour necrosis factor-alpha, interleukin (IL)-1beta, IL-6, IL-8, IL-10 and IL-12) and post-bronchodilator decline in forced expiratory volume in 1 s (FEV(1)) were analysed. PPMs periodically showing the same molecular profile using pulse field gel electrophoresis were considered long-term persistent. Bronchial colonisation was observed in 56 out of 79 follow-up examinations (70.9%) and was mainly due to Haemophilus influenzae, Pseudomonas aeruginosa and enterobacteria (n = 47). These PPMs were all related to sputum neutrophilia (p< or =0.05, Chi-squared test), and H. influenzae was related to higher levels of IL-1beta (p = 0.005) and IL-12 (p = 0.01), with a dose-response relationship (Spearman's correlation coefficient of 0.38 for IL-1beta (p = 0.001), and of 0.32 for IL-12 (p = 0.006)). Haemophilus parainfluenzae was not associated with an identifiable inflammatory response. Long-term persistence of the same strain was observed in 12 examinations (21.4%), mainly due to P. aeruginosa or enterobacteria. A neutrophilic bronchial inflammatory response was associated with a statistically significant decline in FEV(1) during follow-up (OR 2.67, 95% CI 1.07-6.62). A load-related relationship to bronchial inflammation in moderate COPD was observed for colonisation by H. influenzae, but not for colonisation by H. parainfluenzae.

Low-dose theophylline enhances the anti-inflammatory effects of steroids during exacerbations of COPD.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is characterised by an abnormal inflammatory response mainly to cigarette smoke that flares up during exacerbations of the disease (ECOPD). Reduced activity of histone deacetylases (HDAC) contributes to enhanced inflammation in stable COPD. It was hypothesised that HDAC activity is further reduced during ECOPD and that theophylline, an HDAC activator, potentiates the anti-inflammatory effect of steroids in these patients. A study was performed to investigate HDAC activity during ECOPD and the effects of theophylline on the anti-inflammatory effects of steroids in a randomised single-blind controlled study. METHODS: 35 patients hospitalised with ECOPD and treated according to international guidelines (including systemic steroids) were randomised to receive or not to receive low-dose oral theophylline (100 mg twice daily). Before treatment and 3 months after discharge, HDAC and nuclear factor-kappaB (NF-kappaB) activity in sputum macrophages, the concentration of nitric oxide in exhaled air (eNO) and total antioxidant status (TAS), tumour necrosis factor alpha (TNFalpha), interleukin (IL)-6 and IL8 levels in sputum supernatants were measured. RESULTS: Patients receiving standard therapy showed decreased NF-kappaB activity, eNO concentration and sputum levels of TNFalpha, IL6 and IL8, as well as increased TAS during recovery of ECOPD, but HDAC activity did not change. The addition of low-dose theophylline increased HDAC activity and further reduced IL8 and TNFalpha concentrations. CONCLUSIONS: During ECOPD, low-dose theophylline increases HDAC activity and improves the anti-inflammatory effects of steroids. TRIAL REGISTRATION NUMBER: NCT00671151.

Type 1 Diabetes Mellitus reversal via implantation of magnetically purified microencapsulated pseudoislets.

Microencapsulation of pancreatic islets for the treatment of Type I Diabetes Mellitus (T1DM) generates a high quantity of empty microcapsules, resulting in high therapeutic graft volumes that can enhance the host's immune response. We report a 3D printed microfluidic magnetic sorting device for microcapsules purification with the objective to reduce the number of empty microcapsules prior transplantation. In this study, INS1E pseudoislets were microencapsulated within alginate (A) and alginate-poly-L-lysine-alginate (APA) microcapsules and purified through the microfluidic device. APA microcapsules demonstrated higher mechanical integrity and stability than A microcapsules, showing better pseudoislets viability and biological function. Importantly, we obtained a reduction of the graft volume of 77.5% for A microcapsules and 78.6% for APA microcapsules. After subcutaneous implantation of induced diabetic Wistar rats with magnetically purified APA microencapsulated pseudoislets, blood glucose levels were restored into normoglycemia (<200 mg/dL) for almost 17 weeks. In conclusion, our described microfluidic magnetic sorting device represents a great alternative approach for the graft volume reduction of microencapsulated pseudoislets and its application in T1DM disease.

Neonatal pulmonary tuberculosis evolving to a destroyed lung.

Tuberculosis (TB) in the newborn is infrequent, difficult to diagnose and often devastating. Congenital TB is rare, with most neonates and young infants becoming infected after birth. The incidence of neonatal TB might increase in industrialised countries as a result of immigration from countries with higher TB incidence among women of childbearing age. We report two cases of post-natally acquired pulmonary TB in newborns who developed marked lung destruction, a complication of TB which has seldom been described in the first month of life. A high index of clinical suspicion is required when evaluating pregnant women at risk for TB and their ill children, as early identification and treatment can prevent the devastating consequences of TB.

The management of constipation in palliative care: clinical practice recommendations.

Constipation is one of the most common problems in patients receiving palliative care and can cause extreme suffering and discomfort. The aims of this study are to raise awareness of constipation in palliative care, provide clear, practical guidance on management and encourage further research in the area. A pan-European working group of physicians and nurses with significant experience in the management of constipation in palliative care met to evaluate the published evidence and produce these clinical practice recommendations. Four potentially relevant publications were identified, highlighting a lack of clear, practical guidance on the assessment, diagnosis and management of constipation in palliative care patients. Given the limited data available, our recommendations are based on expert clinical opinion, relevant research findings from other settings and best practice from the countries represented. Palliative care patients are at a high risk of constipation, and while general principles of prevention should be followed, pharmacological treatment is often necessary. The combination of a softener and stimulant laxative is generally recommended, and the choice of laxatives should be made on an individual basis. The current evidence base is poor and further research is required on many aspects of the assessment, diagnosis and management of constipation in palliative care.

Concordance between brain 18F-FDG PET and cerebrospinal fluid biomarkers in diagnosing Alzheimer's disease. / Concordancia entre la PET cerebral con 18F-FDG y los biomarcadores en líquido cefalorraquídeo en el diagnóstico de enfermedad de Alzheimer.

OBJECTIVES: Cortical posterior hypometabolism on PET imaging with 18F-FDG (FDG-PET), and altered levels of Aß1-42 peptide, total Tau (tTau) and phosphorylated Tau (pTau) proteins in cerebrospinal fluid (CSF) are established diagnostic biomarkers in Alzheimer's disease (AD). An evaluation has been made of the concordance and relationship between the results of FDG-PET and CSF biomarkers in symptomatic patients with suspected AD. MATERIAL AND METHODS: A retrospective review was carried out on 120 patients with cognitive impairment referred to our Cognitive Neurology Unit, and who were evaluated by brain FDG-PET and a lumbar puncture for CSF biomarkers. In order to calculate their Kappa coefficient of concordance, the result of the FDG-PET and the set of the three CSF biomarkers in each patient was classified as normal, inconclusive, or AD-compatible. The relationship between the results of both methods was further assessed using logistic regression analysis, including the Aß1-42, tTau and pTau levels as quantitative predictors, and the FDG-PET result as the dependent variable. RESULTS: The weighted Kappa coefficient between FDG-PET and CSF biomarkers was 0.46 (95% CI: 0.35-0.57). Logistic regression analysis showed that the Aß1-42 and tTau values together were capable of discriminating an FDG-PET result metabolically suggestive of AD from one non-suggestive of AD, with a 91% sensitivity and 93% specificity at the cut-off line Aß1-42=44+1.3×tTau. CONCLUSIONS: The level of concordance between FDG-PET and CSF biomarkers was moderate, indicating their complementary value in diagnosing AD. The Aß1-42 and tTau levels in CSF help to predict the patient FDG-PET cortical metabolic status.

Silk flow-diverter stent for the treatment of complex intracranial aneurysms: A one-year follow-up multicenter study.

Background Flow-diverter stents have been successfully used in the treatment of complex aneurysms with limited therapeutic alternatives. We report our experience using the Silk flow diverter (SFD; Balt Extrusion, Montmorency, France) for the treatment of complex aneurysms in four Argentine centers. Methods We conducted a retrospective review of 246 consecutive patients who were treated with the SFD at four Argentine centers between January 2009 and January 2017. The patient and aneurysm characteristics, as well as the details of the procedure, were analyzed. The angiographic and clinical findings were recorded during and immediately after the procedure and at 12-month follow-up. Results Angiography follow-up at 12 months was possible in 235 patients (95.5%) with 282 aneurysms. A total of 265 aneurysms (93.9%) presented with complete occlusion of the aneurysmal sac (class 1) and 17 aneurysms (6.1%) presented with partial occlusion (class 2). The 12-month clinical follow-up showed 11 patients with major events (seven, scale 2; five, scale 3; and two, scale 4). The morbidity and mortality rates were 4.2% (11/289) and 2.1% (5/289), respectively. Conclusions The treatment of aneurysms with the SFD was associated with a low rate of complications and a high percentage of aneurysmal occlusion. These findings suggest that SFD is an effective and safe alternative in the endovascular treatment of complex aneurysms.

Motor activity rhythms of forced desynchronized rats subjected to restricted feeding.

Although light is the strongest zeitgeber for the circadian pacemaker, other stimuli can also produce entrainment. In the rat, periodic restricted feeding (RF) is a weak stimulus that may act as a zeitgeber. We tested the effect of RF on the motor activity rhythms of rats subjected to forced dissociation. In this situation two components, supposed to be related with the ventrolateral and dorsomedial areas of the suprachiasmatic nucleus, are detected in their motor activity. One component is entrained to the external light-dark cycle (Light Dependent Component, LDC) and thus has the same period, while the other has a period longer than 24 h (Non-Light Dependent Component, NLDC). This experiment examined whether RF can act on one or both of these two rhythms. Rats were maintained under the light-dark cycles of 22 h (T22) or 23 h (T23) for 44 days with food available for four hours per day. Afterwards the rats received food ad libitum, to test the effect of the previous RF condition. Results show that RF modifies the manifestation of the two initial rhythms, being this effect stronger under T23 than under T22. However RF does not affect the NLDC period. The results reveal that the animal can manifest simultaneously several rhythmic patterns.

Arterial elasticity measurement in renal transplant patients under anticalcineurin immunosuppression.

INTRODUCTION: Calcineurin inhibitors may be associated with decreased arterial elasticity and increased vascular risk. We measured pulse wave velocity (PWV) in large or small arteries as an index of elasticity. The aim of our study was to determine aortic and radial arterial elasticity in 30 stable kidney transplant patients treated with calcineurin inhibitor immunosuppression. PATIENTS AND METHODS: In stable kidney transplant patients we determined the usual biochemical parameters as well as lipid profiles, 24-hour blood pressure (BP) monitoring (CBPM) using a chronobiological program (Garapa), and PWV with a HDI-PWV CR-2000 monitor. RESULTS: Sixteen patients received cyclosporine (CsA, G-1) and 14 tacrolimus (G-2) immunosuppression. There were no baseline differences regarding age (G-1: 56 +/- 12 years, G-2: 56 +/- 14 years), renal transplant follow-up (G-1: 7 +/- 3 years, G-2: 7.5 +/- 3 years), Systolic BP, pulse pressure or plasma creatinine (G-1: 163 +/- 35 umol/L, G-2: 173 +/- 26 umol/L). Patients in the G-1 showed higher diastolic BP (79 +/- 11 vs 74 +/- 8 mm Hg), greater proteinuria (1.26 +/- 0.4 vs 0.6 +/- 0.2 g/d, P < .05), total cholesterol (5.51 +/- 1.2 mmol/L) and low-density lipoprotein (3.08 +/- 0.3 vs 2.99 +/- 0.3 mmol/L, P = NS). Aortic arterial elasticity was decreased in G-1 patients (10.4 +/- 6 vs 14.3 +/- 2 mL/mm Hg x10, P < .05) as well as that in the radial artery (G-1: 5.52 +/- 1 vs 5.57 +/- 1.2 mL/mm Hg x100, P = NS). Almost 100% of the patients presented normal diurnal BP with high nocturnal BP in a nondipper pattern in both groups. CONCLUSION: Calcineurin immunosuppression may contribute to arterial stiffness in kidney transplant patients. No differences between CsA or tacrolimus were observed in our study. CBPM and PWV are useful tools to evaluate subclinical atherosclerosis in renal transplant patients.

Darkness during early postnatal development is required for normal circadian patterns in the adult rat.

Early light experience influences the brain during development. Perinatal light exposure has an important effect on the development of the circadian system, although the role of quantity versus quality of light in this process is still unclear. We tested the development of the circadian rhythm of locomotor activity under constant bright light from the day of weaning, of six groups of rats raised under different light conditions during suckling. Results indicated that when rats received daily darkness during suckling (rats reared under constant darkness or light-dark cycles with dim or bright light) became arrhythmic when exposed to continuous bright light after weaning. However, those rats reared in the absence of darkness (constant dim or bright light, or alternating dim and bright light) developed a circadian rhythm, which was stronger and had a shorter period depending on the quantity of light received during suckling. Vasointestinal polypeptide immunoreactivity in the suprachiasmatic nucleus (SCN) was higher in those rats with weaker rhythms. However, no apparent differences among these groups were found in the melanopsin-expressing retinal ganglion cells, which provide the SCN with light input in the photoentrainment process. When bright light was shifted to dim light in three of the groups on day 57 after weaning, all of them generated a circadian rhythm with a longer period in those rats previously arrhythmic. Our results indicate the importance of the amount of light received at the early stages of life in the development of the circadian system and suggest that darkness is needed for the normal development of circadian behaviour.

Metal exposure influences the melanin and carotenoid-based colorations in great tits.

Metals are naturally found in the environment but are also emitted through anthropogenic activities, raising some concerns about the potential deleterious effects of these elements on wildlife. The potential effects of metals on bird coloration have been the focus of several recent studies since animal colored-signals often reflect the physiology of their bearers and are thus used by animals to assess the quality of another individual as a mate or competitor. These studies have shown that the melanin pigmentation seems to be positively associated and the carotenoid-based coloration negatively associated with metal exposure in wild birds. Although these studies have been very useful to show the associations between metal exposure and coloration, only few of them have actually quantified the levels of metal exposure at the individual level; always focusing on one or two of them. Here, we measured the concentrations of eight metals in great tits' feathers and then assessed how these levels of metals were associated with the carotenoid and melanin-based colorations. We found that the melanin pigmentation was positively associated with the copper concentration and negatively correlated with the chromium concentration in feathers. In addition, we have shown that the carotenoid-based coloration was negatively associated with the feather's mercury concentration. This study is the first one to identify some metals that might affect positively and negatively the deposition of melanin and carotenoid into the plumage of wild birds.

Evolution of rat motor activity circadian rhythm under three different light patterns.

The effects of the light pattern on the evolution of circadian motor activity rhythm of rats were studied in this work. Three different light patterns were used: LL (bright light, 300 lux), DD (dim red light) and LD (12:12 cycles). The animals used for the experiment were born and kept under each condition. At the day of weaning (21-22 days old) animals were isolated and their motor activity was detected by means of an inductive system. Data were recorded every 30 minutes for the first month after weaning. Periodogram analysis was applied to each animal's data and the daily power spectra were calculated on the basis of the endogenous period, tau. The evolution of the rhythm was studied by examining the changes of the whole power spectra of the motor activity function, obtained by means of a Fourier analysis, through time. The power content of the circadian harmonic (PCCH) was considered to be a measure of the circadian character of the function. Results showed the predominancy of ultradian harmonics, when the animals were young, especially in LL, and the increase of the PCCH through time in all cases. Animals under LL show the circadian harmonic to be the main harmonic of the spectra at about day 15 after weaning, while the animals under DD show this harmonic as the main one from the first day. However, the power content of this harmonic increased until day 10. LD animals also showed the first harmonic as the main one from the time of weaning increasing the PCCH until day 7. These results are explained in respect to a multioscillatory system.(ABSTRACT TRUNCATED AT 250 WORDS)

Sex differences in the development of the motor activity circadian rhythm in rats under constant light.

Differences between sexes in the maturation of the motor activity rhythm have been studied in rats (10 females and 7 males) born and raised under LL and isolated from the day of weaning. Motor activity data were registered every 30 minutes for 45 days after weaning. Periodogram analysis was applied to each animal data and the daily power spectra were calculated on the basis of the period, tau. The process of rhythm evolution was studied by means of an analysis of variance of a multivariant linear model, and by defining two mathematical functions that fit to the evolution of the total daily motor activity (TDMA) and the power content of the circadian harmonic (PCCH) through time: a sigmoidal function, which presents an inflexion was used for the study of the evolution of the PCCH values through time: and a gaussian function, which presents a minimum was used for the evolution of the TDMA values. Results indicate that males show more ultradian components than females just after weaning, and take a longer time to acquire the circadian harmonic as the main harmonic in the daily power spectrum. The acquisition of the circadian character is preceded by a decrease in the TDMA, in the case of the males and is followed by a decrease in TDMA in the case of the females. These results suggest sexual differences in the organization and maturation of the circadian system in rats.

Sound does not entrain the motor activity circadian rhythm of rats.

The effect of sound/silence cycles on the motor activity rhythm of the rat was studied. The daily motor activity pattern was studied in two groups of rats of both sexes kept first under DD and later under LL. One group (16 rats) was subjected to a cycle of sound and silence and the other (8 rats) was used as control. The study was performed under two different light regimes, to consider the possibility that a weak zeitgeber (such as sound might be), would exert an effect on the circadian pacemaker. This effect would depend on the stability of the circadian system, which could be modified by the lighting conditions. Results show no statistically significant differences between the two groups, neither in the motor activity patterns nor in the period of the rhythm. It was concluded that sound is not a zeitgeber for the motor activity rhythm of rats.

Hereditary nature of the pattern of the motor activity circadian rhythm in mice.

The inherited nature of the pattern of the daily motor activity function (MAF) has been studied using different families of the same strain of mice. The motor activity of 6 male mice, belonging to two different families (A and B was recorded during 54 days in LD conditions (12L:12D) using six inductive units. The register began the day just after weaning. A Fourier analysis was applied to motor activity data corresponding to observation periods of 24 hours. The daily power spectrum, with 20 harmonics, was calculated by obtaining the power content of each harmonic expressed as a percentage of the total power content of the MAF. A discriminant function based on the first fifteen harmonics was created to distinguish between the males of the two families. Then, the same discriminant function was applied to the daily power spectra of the progeny. It was also possible to differentiate among the mice (males and females) coming from fathers of family A and the mice coming from fathers of family B (p less than 0.02). This fact suggests that, in the power spectra of the progeny, there may be enough characteristic equal to that of their respective fathers and it further suggests that some characteristic of the MAF could have been inherited.

Dissociation of the rat motor activity rhythm under T cycles shorter than 24 hours.

Since the suprachiasmatic nuclei (SCN) were identified as the principal mammalian circadian clock, many studies describing their morphology and physiology have been carried out. Today, the multioscillatory nature of the SCN, which explains the dissociation of the circadian rhythms under some experimental conditions, is widely accepted. Here, we study the simultaneous presence of two circadian rhythms in the motor activity of the rat when exposed to symmetric light-dark (LD) cycles shorter than 24 h (T21, T21.5, T22, T22.5, T23, and T23.5). One rhythmic component was entrained by the external LD cycle whereas the other ran free with a period longer than 24 h. The results show that two circadian rhythms were present only when T was shorter than T23, whereas at T23.5 only one entrained component was manifested. The manifestation of the two circadian components depends quantitatively on the period of the external cycle--i.e., the strength of the entrained rhythm increases when the external T is closer to 24 h--whereas that of the nonentrained rhythm decreases. The dissociation of the motor activity rhythm and the gradual appearance of the two components are explained by considering the entrainment of a multioscillatory system as not taking place as a whole but rather in a partial manner, in such a way that some oscillators may entrain but not others. The effect of the entrained oscillators is added to the masking effect of the LD cycles.

Entrainment of the rat motor activity rhythm: effects of the light-dark cycle and physical exercise.

The circadian system is believed to be composed of a population of oscillators that couple together and generate a single rhythm. If this coupling is not strong enough, the circadian system can be dissociated into two or more groups of oscillators, and this is manifested in a dissociation of the overt rhythm into at least two circadian components. This study aims to examine the influence of factors, such as the difference in impact between T and tau, light intensity, and access to a running wheel, on the distribution of motor activity throughout the light-dark (LD) cycle and the dissociation of the rhythm. Rats were submitted to LD cycles of 23 h (T23) or 25 h. For each such cycle, half the rats were submitted to high light intensity and the other half to low light intensity. For each of these conditions, half the rats were kept in small cages, and the other half were in cages with a running wheel. Rats were maintained first under LD cycles and afterwards under constant darkness (DD). Motor activity was recorded throughout the whole experiment by means of activity meters with infrared beams. Results show that the distribution of motor activity throughout the cycle and the after effects observed in the rhythm under DD depended on light intensity and access to the wheel. Moreover, under T23, some rats showed two simultaneous circadian components whose manifestation also depended on the experimental conditions. The results indicate that the strength of circadian entrainment to LD cycles in the rat depends on three factors: the period length of the LD cycle, light intensity used during the light phase, and access to a running wheel.

Influence of period length of light/dark cycles on the body weight and food intake of young rats.

The purpose of this work is to test whether physiological factors, such as body weight or food intake, are related to the period of environmental cycles under which animals are kept. We studied body weight increase, daily food intake and efficiency (i.e. relationship between body weight increase and food intake) of 7 groups each containing 20-24 male rats. Measurements were recorded every 4 days from the day of weaning (25 days old) during 64 days. Each group of rats was kept under a symmetrical LD cycle. The period of the cycles studied were: 22, 23, 24, 25, 26, 27 and 28 hours (T22, T23 ... T28). Results show differences in the three variables studied for groups T25 and T26. These animals show an inferior body weight, less food intake and less efficiency. It would seem that the difference between the period of the external LD cycle and the endogenous period of the animals rhythm induces changes in the energetic needs of the organism.