An Enzyme Immunoassay for Determining Immunoreactive Trypsinogen (IRT) in Dried Blood Spots on Filter Paper Using an Ultra-Microanalytical System. Castells Martínez EM, González EC, Tejeda Y, Frómeta A, Martín O, Espinosa M, et al. Appl Biochem Biotechnol. 2018 May 29. DOI: 10.1007/s12010-018-2785-4. [Epub ahead of print]

Cystic fibrosis (CF) is a severe autosomal recessive disorder. It is caused by mutations in the CF transmembrane conductance regulator gene. Early diagnosis of CF can be carried out by determining high immunoreactive trypsinogen (IRT) blood values in newborns. A simple sandwich-type ultramicroELISA assay (UMELISA®) has been developed for the measurement of IRT in dried blood spots on filter paper. Strips coated with a high affinity monoclonal antibody directed against IRT are used as solid phase, to ensure the specificity of the assay. The assay is carried out within 20 h. The useful rank of the curve is 0-500 ng/mL, and the lowest detectable concentration is 4.8 ng/mL. Intra- and inter-assay coefficients of variation were lower than 10%. The recovery mean value was 100.3 ± 11.2%. Cross-reactivity with proteins structurally related to IRT (α2-macroglobulin, α1-antitrypsin, and human chymotrypsin) was lower than the detection limit of the assay. Four thousand four hundred six newborn samples from the Cuban Newborn Screening Program were analyzed, and the mean IRT concentration was 12.8 ng/mL. Higher IRT values were obtained when samples were eluted overnight. Regression analysis showed a good correlation with the commercially available AutoDELFIA® Neonatal IRT kit (n = 3948, r = 0.885, ƙ = 0.976, p < 0.01). The analytical performance characteristics of our UMELISA® TIR Neonatal suggest that it can be used for the neonatal screening of CF.

APACHE II score for critically ill patients with a solid tumor: A reclassification study. Martos-Benítez FD, Cordero-Escobar I, Soto-García A, Betancourt-Plaza I, González-Martínez I. Rev Esp Anestesiol Reanim. 2018 May 17. pii: S0034-9356(18)30092-6.
DOI: 10.1016/j.redar.2018.04.001. [Epub ahead of print]

Objective To improve the accuracy of the Acute Physiology and Chronic Health Evaluation (APACHE) II model for predicting hospital mortality in critically ill cancer patients. Materials and Methods This was a prospective cohort study of 522 patients admitted to ICU with a solid tumor. We developed the “APACHE II score for critically ill patients with a solid tumor” (APACHE II_CCP score), in which typical variables of critically ill cancer patients were added to general APACHE II score. Calibration and discrimination were evaluated by Hosmer-Lemeshow test (H-L) and area under receiver operating characteristic curve (AROC), respectively. The improvement in predicting hospital mortality with the new model was assessed using a reclassification analysis by integrated discrimination improvement (IDI), net reclassification improvement (NRI; cut-off point of 20% in risk of death) and quantitative NRI (qNRI). Results The hospital mortality rate was 13%. Discrimination was superior for APACHE II_CCP score (AROC=0.91 [95% CI 0.87-0.94; P<.0001]) compared to general APACHE II score (AROC=0.62 [95% CI 0.54-0.70; P=.002]). Calibration was better using APACHE II_CCPscore (H-L; P=.267 vs. P=.001). In reclassification analysis, an improved mortality prediction was observed with APACHE II_CCP score (IDI=0.2994 [P<.0001]; total qNRI=134.3% [95% CI 108.8-159.8%; P<.0001]; total NRI=41.5% [95% CI 23.7-59.3%; P<.0001]). Conclusions The performance of APACHE II_CCP score was superior to that observed for general APACHE II score in predicting mortality in critically ill patients with a solid tumor. Other studies validating this new predictive model are required.

A physical model of cell metabolism. Fernández-de-Cossio-Díaz J, Vázquez A. Sci Rep. 2018 May 29;8(1):8349. DOI: 10.1038/s41598-018-26724-7.

Cell metabolism is characterized by three fundamental energy demands: to sustain cell maintenance, to trigger aerobic fermentation and to achieve maximum metabolic rate. The transition to aerobic fermentation and the maximum metabolic rate are currently understood based on enzymatic cost constraints. Yet, we are lacking a theory explaining the maintenance energy demand. Here we report a physical model of cell metabolism that explains the origin of these three energy scales. Our key hypothesis is that the maintenance energy demand is rooted on the energy expended by molecular motors to fluidize the cytoplasm and counteract molecular crowding. Using this model and independent parameter estimates we make predictions for the three energy scales that are in quantitative agreement with experimental values. The model also recapitulates the dependencies of cell growth with extracellular osmolarity and temperature. This theory brings together biophysics and cell biology in a tractable model that can be applied to understand key principles of cell metabolism.

Contrasting Timing of Virological Relapse after Discontinuation of Tenofovir or Entecavir in Hepatitis B e Antigen-negative Patients. Höner Zu Siederdissen C, Hui AJ, Sukeepaisarnjaroen W, Tangkijvanich P, Su WW, Nieto GEG, et al. J Infect Dis. 2018 Jun 9. DOI: 10.1093/infdis/jiy350. [Epub ahead of print]

Stopping long-term nucleos(t)ide analogue therapy increases HBsAg loss rates in HBeAg-negative patients. Viral rebound may induce immune responses facilitating functional cure. We analyzed which factors are associated with timing of virological relapse in 220 Asian HBeAg-negative patients from the prospective ABX203 vaccine study. Unexpectedly, only the type of antiviral therapy was significantly associated with early virological relapse, defined as HBV DNA > 2,000 IU/ml until week 12 and occurred earlier in patients treated with tenofovir versus entecavir (median time 6 versus 24 weeks, p < 0.0001). This should be considered for future trials and monitoring of patients after treatment discontinuation.

Dose-response study for the highly aggressive and metastatic primary F3II mammary carcinoma under direct current. González MM, Morales DF, Cabrales LEB, Pérez DJ, Montijano JI, Castañeda ARS, et al. Bioelectromagnetics. 2018 Jun 5.
DOI: 10.1002/bem.22132. [Epub ahead of print]

Electrochemical treatment has been suggested as an effective alternative to local cancer therapy. Nevertheless, its effectiveness decreases when highly aggressive primary tumors are treated. The aim of this research was to understand the growth kinetics of the highly aggressive and metastatic primary F3II tumor growing in male and female BALB/c/Cenp mice under electrochemical treatment. Different amounts of electric charge (6, 9, and 18 C) were used. Two electrodes were inserted into the base, perpendicular to the tumor’s long axis, keeping about 1 cm distance between them. Results have shown that the F3II tumor is highly sensitive to direct current. The overall effectiveness (complete response + partial response) of this physical agent was ≥75.0% and observed in 59.3% (16/27) of treated F3II tumors. Complete remission of treated tumors was observed in 22.2% (6/27). An unexpected result was the death of 11 direct current-treated animals (eight females and three males). It is concluded that direct current may be addressed to significantly affect highly aggressive and metastatic primary tumor growth kinetics, including the tumor complete response.

Efficacy of Four Solanum spp. Extracts in an Animal Model of Cutaneous Leishmaniasis. Cos P, Janssens J, Piñón A, Cuesta-Rubio O, Yglesias-Rivera A, Díaz-García A, et al. Medicines (Basel). 2018 Jun 5;5(2). pii: E49. DOI: 10.3390/medicines5020049.

Background Leishmaniasis is a complex protozoa disease caused by Leishmania genus (Trypanosomatidae family). Currently, there have been renewed interests worldwide in plants as pharmaceutical agents. In this study, the in vivo efficacy of Solanum spp. is assessed in an L. amazonensis BALB/c mice model for experimental cutaneous leishmaniasis. Methods Animals were infected with 5 × 10⁶ metacyclic promastigotes and 30-day post-infection, a treatment with 30 mg/kg of Solanum extracts or Glucantime^® (GTM) was applied intralesionally every four days to complete 5 doses. Results Neither death nor loss of weight higher than 10% was observed. All the tested extracts were able to control the infection, compared with the infected and untreated group. Solanum havanense Jacq. extract showed the highest efficacy and was superior (p < 0.05) to GTM. Solanum myriacanthum Dunal., S. nudum Dunal. and S. seaforthianum Andr. extracts demonstrated a similar effect (p > 0.05) to GTM. An increase of IFN-γ (p < 0.05) was displayed only by animals treated with S. nudum compared to the group treated with a vehicle, while no differences (p > 0.05) were observed for IL-12. Conclusions In vivo effects of Solanum extracts were demonstrated, suggesting that this genus could be further explored as a new antileishmanial alternative.

Epilepsy mortality trends in Cuba compared with England and Wales: 1987-2010. Suárez-Medina R, Morales-Chacón LM, Venero-Fernández SJ, Liu C, Fogarty AW, Neligan A. Epilepsy Behav. 2018 Jun 13;85:72–5. DOI: 10.1016/j.yebeh.2018.04.031. [Epub ahead of print]

There are limited epilepsy mortality data from developing countries and Latin America in particular. We examined national epilepsy mortality data from Cuba and contrasted them with comparable data from England and Wales. National epilepsy mortality data for Cuba between the years 1987 and 2010 were obtained from the Medical Records and Health Statistics Bureau of the Cuban Public Health Ministry (www.sld.cu/sitios/dne/) with the corresponding mortality data from England and Wales obtained from the UK Office of National Statistics (ONS, www.ons.gov.uk). Indirect standardization with calculation of a standardized mortality ratio (SMR) was used to compare trends. The overall trend was of a slight decrease in mortality rates over the 23 years in Cuba, with higher mortality rates primarily occurring in young people. Annual age-adjusted rates were consistently lower in Cuba than those seen in England and Wales, with the SMR ranging from 0.35 (95% confidence interval (CI): 0.30 to 0.48) in 2007 to 1.00 (95% CI: 0.85 to 1.15) in 1994. Cuban epilepsy mortality rates are consistently lower than those of England and Wales. Reasons for this disparity in mortality rates are not immediately apparent but are likely to be multifactorial.

Factors associated with ATXN2 CAG/CAA repeat intergenerational instability in Spinocerebellar Ataxia type 2. Almaguer-Mederos LE, Mesa JML, González-Zaldívar Y, Almaguer-Gotay D, Cuello-Almarales D, Aguilera-Rodríguez R, et al. Clin Genet. 2018 May 14. DOI: 10.1111/cge.13380. [Epub ahead of print]

Spinocerebellar ataxia type 2 (SCA2) is a neurodegenerative disorder caused by the unstable expansion of a CAG/CAA repeat in the ATXN2 gene, which normally encodes 22 glutamines (Q22). A large study was conducted to characterize the CAG/CAA repeat intergenerational instability in SCA2 families. Large normal alleles (LNA, Q24-31) were significantly more unstable upon maternal transmissions. In contrast, expanded alleles (EA, Q32-750) were significantly more unstable during paternal transmissions, in correlation with repeat length. Significant correlations were found between the instability and the age at conception in paternal transmissions. In conclusion, intergenerational instability at ATXN2 locus is influenced by the sex, repeat length and age at conception of the transmitting parent. These results have profound implications for genetic counseling services.

Fibrosis Severity as a Determinant of Cause-specific Mortality in Patients With Advanced Nonalcoholic Fatty Liver Disease. Vilar-Gómez E, Calzadilla-Bertot L, Wai-Sun Wong V, Castellanos M, Aller-de la Fuente R, Metwally M, et al. Gastroenterology. 2018 May 4.
pii: S0016-5085(18)34484-6. DOI: 10.1053/j.gastro.2018.04.034. [Epub ahead of print]

Background & Aims Little is known about the natural course of nonalcoholic fatty liver disease (NAFLD) with advanced fibrosis. We describe long-term outcomes and evaluate the effects of clinical and histologic parameters on disease progression in patients with advanced NAFLD. Methods We conducted a multi-national study of 458 patients with biopsy-confirmed NAFLD with bridging fibrosis (F3, n=159) or compensated cirrhosis (222 patients with Child-Turcotte-Pugh [CTP] scores of A5 and 77 patients with scores of A6), evaluated from April 1995 through November 2013 and followed until December 2016, death, or liver transplantation at hepatology centers in Spain, Australia, Hong Kong, and Cuba. Biopsies were reevaluated and scored; demographic, clinical, laboratory, and pathology data for each patient were collected from the time of liver biopsy collection. Cox proportional and competing risk models were used to estimate rates of transplant-free survival and major clinical events and to identify factors associated with outcomes. Results During a mean follow-up time of 5.5 years (range, 2.7-8.2 years), 37 patients died, 37 received liver transplants, 88 had initial hepatic decompensation events, 41 developed hepatocellular carcinoma (HCC), 14 had vascular events, and 30 developed non-hepatic cancers. A higher proportion of patients with F3 fibrosis survived transplant-free for 10 years (94%; 95% CI, 86-99) than of patients with cirrhosis and CTP-A5 (74%; 95% CI, 61-89) or CTP-A6 (17%; 95% CI, 6-29). Patients with cirrhosis were more likely than patients with F3 fibrosis to have hepatic decompensation (44%; 95% CI, 32-60 vs 6%, 95% CI, 2-13) or HCC (17%; 95% CI, 8-31 vs 2.3%, 95% CI, 1-12). The cumulative incidence of vascular events was higher in patients with F3 fibrosis (7%; 95% CI, 3-18) than cirrhosis (2%; 95% CI, 0-6). The cumulative incidence of non-hepatic malignancies was higher in patients with F3 fibrosis (4%; 95% CI, 7-23) than cirrhosis (6%; 95% CI, 2-15). Death or transplantation, decompensation, and HCC were independently associated with baseline cirrhosis and mild (<33%) steatosis whereas moderate alcohol consumption associated with these outcomes only in patients with cirrhosis. Conclusions Patients with NAFLD cirrhosis have predominantly liver-related events whereas those with bridging fibrosis have predominantly non-hepatic cancers and vascular events.

Giardiasis: a diagnosis that should be considered regardless of the setting. Escobedo AA, Almirall P, Hanevik K, Cimerman S, Rodríguez-Morales AJ, Almanza C, et al. Epidemiol Infect. 2018 Jun 11:1–3. DOI: 10.1017/S0950268818001504. [Epub ahead of print]

Although Giardia, the aetiological agent of giardiasis, is one of the most prevalent intestinal parasitic infections world-wide, for industrialised countries, it is mainly appreciated as an imported disease with the minimal local transmission. However, the current evidence challenges this perception; Giardia has relevance beyond the high prevalence areas. This infection may be asymptomatic or cause gastrointestinal complains and long-term sequelae, including irritable bowel syndrome, chronic fatigue and impaired child growth and cognitive development. Its detection and diagnosis present a challenge to physicians who may not be familiar with this infection. To improve interventions to control this parasitosis, it is necessary to maintain a high index of suspicion and remain vigilant in finding cases at risk for infection. A better understanding of the characteristics of populations importing infections alongside improved methods to reliably classify infections as imported or acquired locally will help to ensure early and accurate diagnosis. The evidence shows that public health problems like giardiasis are global issues that need to be addressed collectively by both high and low prevalence countries.

JM-20 Treatment After MCAO Reduced Astrocyte Reactivity and Neuronal Death on Peri-infarct Regions of the Rat Brain. Ramírez-Sánchez J, Pires ENS, Meneghetti A, Hansel G, Nuñez-Figueredo Y, Pardo-Andreu GL, et al. Mol Neurobiol. 2018 May 3.
DOI: 10.1007/s12035-018-1087-8. [Epub ahead of print]

Stroke is frequently associated with severe neurological decline and mortality, and its incidence is expected to increase due to aging population. The only available pharmacological treatment for cerebral ischemia is thrombolysis, with narrow therapeutic windows. Efforts aimed to identify new therapeutics are crucial. In this study, we look into plausible molecular and cellular targets for JM-20, a new hybrid molecule, against ischemic stroke in vivo. Male Wistar rats were subjected to 90 min middle cerebral artery occlusion (MCAO) following 23 h of reperfusion. Animals treated with 8 mg/kg JM-20 (p.o., 1 h after reperfusion) showed minimal neurological impairment and lower GABA and IL-1β levels in CSF when compared to damaged rats that received vehicle. Immunocontent of pro-survival, phosphorylated Akt protein decreased in the cortex after 24 h as result of the ischemic insult, accompanied by decreased number of NeuN⁺ cells in the peri-infarct cortex, cornu ammonis 1 (CA1) and dentate gyrus (DG) areas. Widespread reactive astrogliosis in both cortex and hippocampus (CA1, CA3, and DG areas) was observed 24 h post-ischemia. JM-20 prevented the activated Akt reduction, neuronal death, and astrocytes reactivity throughout the brain. Overall, the results reinforce the pharmacological potential of JM-20 as neuroprotective agent and provide important evidences about its molecular and cellular targets in this model of cerebral ischemia.

Junior temperament character inventory together with quantitative EEG discriminate children with attention deficit hyperactivity disorder combined subtype from children with attention deficit hyperactivity disorder combined subtype plus oppositional defiant disorder. Chiarenza GA, Villa S, Galán L, Valdés-Sosa P, Bosch-Bayard J. Int J Psychophysiol. 2018 May 19;130:9–20. DOI: 10.1016/j.ijpsycho.2018.05.007. [Epub ahead of print]

Oppositional defiant disorder (ODD) is frequently associated with Attention Deficit Hyperactivity Disorder (ADHD) but no clear neurophysiological evidence exists that distinguishes the two groups. Our aim was to identify biomarkers that distinguish children with Attention Deficit Hyperactivity Disorder combined subtype (ADHD_C) from children with ADHD_C + ODD, by combining the results of quantitative EEG (qEEG) and the Junior Temperament Character Inventory (JTCI). 28 ADHD_C and 22 ADHD_C + ODD children who met the DSMV criteria participated in the study. JTCI and EEG were analyzed. Stability based Biomarkers identification methodology was applied to the JTCI and the qEEG separately and combined. The qEEG was tested at the scalp and the sources levels. The classification power of the selected biomarkers was tested with a robust ROC technique. The best discriminant power was obtained when TCI and qEEG were analyzed together. Novelty seeking, self-directedness and cooperativeness were selected as biomarkers together with F4 and Cz in Delta; Fz and F4 in Theta and F7 and F8 in Beta, with a robust AUC of 0.95 for the ROC. At sources level: the regions were the right lateral and medial orbito-frontal cortex, cingular region, angular gyrus, right inferior occipital gyrus, occipital pole and the left insula in Theta, Alpha and Beta. The robust estimate of the total AUC was 0.91. These structures are part of extensive networks of novelty seeking, self-directedness and cooperativeness systems that seem dysregulated in these children. These methods represent an original approach to associate differences of personality and behavior to specific neuronal systems and subsystems.

Knowledge and beliefs about dementia among the general public: A preliminary report on the Cuban population. Broche-Pérez Y, Fernández-Fleites Z, González B, Hernández Pérez MA, Salazar-Guerra YI. Neurologia. 2018 May 8. pii: S0213-4853(18)30098-7.
DOI: 10.1016/j.nrl.2018.03.011. [Epub ahead of print]

Background The number of people diagnosed with dementia globally has dramatically increased in recent years. The objective of this study was to explore beliefs and knowledge among the Cuban population with regard to the risk factors that may lead to dementia and the actions that may be taken to prevent it. Methods In an exploratory cross-sectional study, we surveyed a total of 391 people aged between 18 and 96 years. The results were stratified by sex, age range, level of education, and contact with dementia. Results Dementia was the fourth most worrying disease. A total of 64.5% of participants believed that the risk of dementia could be reduced, and 60% that the appropriate time to begin prevention measures is after the age of 40. Cognitive stimulation and healthy diet were more frequently cited as useful activities to reduce risk. Survey respondents reported little presence in their lifestyle of behaviours that are beneficial for reducing the risk of dementia. Conclusions Although dementia is an important health issue for respondents, their knowledge about disease prevention is still insufficient. The results obtained constitute a starting point for the design of policies aimed at increasing knowledge about the disease and improving prevention.

Multi-subject hierarchical inverse covariance modelling improves estimation of functional brain networks. Colclough GL, Woolrich MW, Harrison SJ, Rojas López PA, Valdés-Sosa PA, Smith SM. Neuroimage. 2018 May 7. pii: S1053-8119(18)30397-5.
DOI: 10.1016/j.neuroimage.2018.04.077. [Epub ahead of print]

A Bayesian model for sparse, hierarchical inverse covariance estimation is presented, and applied to multi-subject functional connectivity estimation in the human brain. It enables simultaneous inference of the strength of connectivity between brain regions at both subject and population level, and is applicable to fmri, meg and eeg data. Two versions of the model can encourage sparse connectivity, either using continuous priors to suppress irrelevant connections, or using an explicit description of the network structure to estimate the connection probability between each pair of regions. A large evaluation of this model, and thirteen methods that represent the state of the art of inverse covariance modelling, is conducted using both simulated and resting-state functional imaging datasets. Our novel Bayesian approach has similar performance to the best extant alternative, Ng et al.’s Sparse [Epub ahead of print] Group Gaussian Graphical Model algorithm, which also is based on a hierarchical structure. Using data from the Human Connectome Project, we show that these hierarchical models are able to reduce the measurement error in meg beta-band functional networks by 10%, producing concomitant increases in estimates of the genetic influence on functional connectivity.

Neuroimmunology Research. A Report from the Cuban Network of Neuroimmunology. Robinson-Agramonte MLA, Pedre LL, Serrano-Barrera OR. Behav Sci (Basel). 2018 May 8;8(5). pii: E47. DOI: 10.3390/bs8050047

Neuroimmunology can be traced back to the XIX century through the descriptions of some of the disease’s models (e.g., multiple sclerosis and Guillain Barret syndrome, amongst others). The diagnostic tools are based in the cerebrospinal fluid (CSF) analysis developed by Quincke or in the development of neuroimmunotherapy with the earlier expression in Pasteur’s vaccine for rabies. Nevertheless, this field, which began to become delineated as an independent research area in the 1940s, has evolved as an innovative and integrative field at the shared edges of neurosciences, immunology, and related clinical and research areas, which are currently becoming a major concern for neuroscience and indeed for all of the scientific community linked to it. The workshop focused on several topics: (1) the molecular mechanisms of immunoregulation in health and neurological diseases, (like multiple sclerosis, autism, ataxias, epilepsy, Alzheimer and Parkinson’s disease); (2) the use of animal models for neurodegenerative diseases (ataxia, fronto-temporal dementia/amyotrophic lateral sclerosis, ataxia-telangiectasia); (3) the results of new interventional technologies in neurology, with a special interest in the implementation of surgical techniques and the management of drug-resistant temporal lobe epilepsy; (4) the use of non-invasive brain stimulation in neurodevelopmental disorders; as well as (5) the efficacy of neuroprotective molecules in neurodegenerative diseases. This paper summarizes the highlights of the symposium.

Oxidative Stress in Patients with Drug Resistant Partial Complex Seizure. Lorigados Pedre L, Gallardo JM, Morales Chacón LM, Vega García A, Flores-Mendoza M, Neri-Gómez T, et al. Behav Sci (Basel). 2018 Jun 9;8(6). pii: E59. DOI: 10.3390/bs8060059.

Oxidative stress (OS) has been implicated as a pathophysiological mechanism of drug-resistant epilepsy, but little is known about the relationship between OS markers and clinical parameters, such as the number of drugs, age onset of seizure and frequency of seizures per month. The current study’s aim was to evaluate several oxidative stress markers and antioxidants in 18 drug-resistant partial complex seizure (DRPCS) patients compared to a control group (age and sex matched), and the results were related to clinical variables. We examined malondialdehyde (MDA), advanced oxidation protein products (AOPP), advanced glycation end products (AGEs), nitric oxide (NO), uric acid, superoxide dismutase (SOD), glutathione, vitamin C, 4-hydroxy-2-nonenal (4-HNE) and nitrotyrosine (3-NT). All markers except 4-HNE and 3-NT were studied by spectrophotometry. The expressions of 4-HNE and 3-NT were evaluated by Western blot analysis. MDA levels in patients were significantly increased (p ≤ 0.0001) while AOPP levels were similar to the control group. AGEs, NO and uric acid concentrations were significantly decreased (p ≤ 0.004, p ≤ 0.005, p ≤ 0.0001, respectively). Expressions of 3-NT and 4-HNE were increased (p ≤ 0.005) similarly to SOD activity (p = 0.0001), whereas vitamin C was considerably diminished (p = 0.0001). Glutathione levels were similar to the control group. There was a positive correlation between NO and MDA with the number of drugs. The expression of 3-NT was positively related with the frequency of seizures per month. There was a negative relationship between MDA and age at onset of seizures, as well as vitamin C with seizure frequency/month. We detected an imbalance in the redox state in patients with DRCPS, supporting oxidative stress as a relevant mechanism in this pathology. Thus, it is apparent that some oxidant and antioxidant parameters are closely linked with clinical variables.

Palatal Position of Patient Tracker for Magnetic Neuronavigation System: Technical Note. Catapano G, Sgulò FG, Acurio Padilla PE, Spennato P, Di Nuzzo G, Boniello V, et al. World Neurosurg. 2018 May 9. pii: S1878-8750(18)30942-2. DOI: 10.1016/j.wneu.2018.04.221. [Epub ahead of print]

Objective Recently, the neuronavigation system (NS) has become an essential intraoperative tool for many neurosurgical procedures allowing for precise lesion localization. For such reason, it is particularly important to avoid errors during the navigation process. In the present study focus on a novel technique with the palatal position of patient tracker to ensure an optimal accuracy during magnetic navigation in different neurosurgical procedures. Methods The present retrospective study included a total of 34 patients treated in our Institution between June 2017 and January 2018. Surgical procedures included two groups operated under general anesthesia: microscopic transcranial approach and endoscopic endonasal approach. Preoperative and post-operative navigation accuracy was assessed by two different neurosurgeons. Results After our surgical planning navigation protocol was applied, both transcranial and endonasal procedures were successfully performed under navigation guidance in all patients except one. Intraoperative and post-operative complications related to tracker mounted under the hard palate did not occur. In 33 cases a maximal tracking view and optimal navigation accuracy was achieved accounting for a successful rate of 97%. Conclusions The position of the patient tracker under the hard palate showed safety, accuracy and feasibility in 97% of our patient. In our case series, it allowed to obtain the main goal to avoid device displacement without invasiveness and postoperative discomfort of the patient.

Presence of the p.L456V polymorphism in Cuban patients clinically diagnosed with Wilson’s disease. Clark-Feoktistova Y, Ruenes-Domech C, García-Bacallao EF, Roblejo-Balbuena H, Feoktistova L, Clark-Feoktistova I, et al. Rev Gastroenterol Mex. 2018 Jun 10. pii: S0375-0906(18)30094-6. DOI: 10.1016/j.rgmx.2018.03.005. [Epub ahead of print]

Introduction and Aims Wilson’s disease is characterized by the accumulation of copper in different organs, mainly affecting the liver, brain, and cornea, and is caused by mutations in the ATP7B gene. More than 120 polymorphisms in the ATP7B gene have been reported in the medical literature. The aim of the present study was to identify the conformational changes in the exon 3 region of the ATP7B gene and detect the p.L456V polymorphism in Cuban patients clinically diagnosed with Wilson’s disease. Materials and Methods A descriptive study was conducted at the Centro Nacional de Genética Médica and the Instituto Nacional de Gastroenterología within the time frame of 2007-2012 and included 105 patients with a clinical diagnosis of Wilson’s disease. DNA extraction was performed through the salting-out method and the fragment of interest was amplified using the polymerase chain reaction technique. The conformational shift changes in the exon 3 region and the presence of the p.L456V polymorphism were identified through the Single-Strand Conformation Polymorphism analysis. Results The so-called b and c conformational shift changes, corresponding to the p.L456V polymorphism in the heterozygous and homozygous states, respectively, were identified. The allelic frequency of the p.L456V polymorphism in the 105 Cuban patients that had a clinical diagnosis of Wilson’s disease was 41% and liver-related symptoms were the most frequent in the patients with that polymorphism. Conclusion The p.L456V polymorphism was identified in 64 Cuban patients clinically diagnosed with Wilson’s disease, making future molecular study through indirect methods possible.

Protective Activity of Erythropoyetine in the Cognition of Patients with Parkinson’s Disease. Pedroso I, García M, Casabona E, Morales L, Bringas ML, Pérez L, et al. Behav Sci (Basel). 2018 May 21;8(5):51. DOI: 10.3390/bs8050051. eCollection 2018 May.

Introduction Treatment strategies in Parkinson’s disease (PD) can improve a patient’s quality of life but cannot stop the progression of PD. We are looking for different alternatives that modify the natural course of the disease and recent research has demonstrated the neuroprotective properties of erythropoietin. In Cuba, the Center for Molecular Immunology (CIM) is a cutting edge scientific center where the recombinant form (EPOrh) and recombinant human erythropoietin with low sialic acid (NeuroEPO) are produced. We performed two clinical trials to evaluate the safety and tolerability of these two drugs in PD patients. In this paper we want to show the positive results of the additional cognitive tests employed, as part of the comprehensive assessment. Materials and Methods Two studies were conducted in PD patients from the outpatient clinic of CIREN, including n = 10 and n = 26 patients between 60 and 66 years of age, in stages 1 to 2 of the Hoehn and Yahr Scale. The first study employed recombinant human (rhEPO) and the second an intranasal formulation of neuroEPO. All patients were evaluated with a battery of neuropsychological scales composed to evaluate global cognitive functioning, executive function, and memory. Results The general results in both studies showed a positive response to the cognitive functions in PD patients, who were undergoing pharmacological treatment with respect to the evaluation (p < 0.05) before the intervention. Conclusions Erythropoietin has a discrete positive effect on the cognitive functions of patients with Parkinson’s disease, which could be interpreted as an effect of the neuroprotective properties of this molecule. To confirm the results another clinical trial phase III with neuroEPO is in progress, also designed to discard any influence of a placebo effect on cognition.

Quantitative ELISA sandwich for a new vaccine against avian influenza virus H5N1. Pose AG, Rodríguez ER, Piñeiro MJ, Montesino R, Sánchez O, Toledo JR. J Immunol Methods. 2018 May 24. pii: S0022-1759(17)30554-9. DOI: 10.1016/j.jim.2018.05.008. [Epub ahead of print]

Analytical techniques are essential in the process of standardizing and validating vaccines. In this study we described a methodology to establish an ELISA sandwich for the quantification of a new vaccine against avian influenza virus H5N1 based on the main antigenic determinant of the virus, the extracellular domain of the glycoprotein hemagglutinin (HA), fused to the extracellular domain of the chicken CD154 glycoprotein (HACD). The chimerical proteins HA and HACD were produced in SiHa cells and the experiments were performed by using three monoclonal antibodies (MAb-HA1, MAb-HA2 and MAb-HA3), alone or conjugated to horseradish peroxidase (HRP-HA1, HRP-HA2 and HRP-HA3). The hemagglutination inhibition assay was carried out with a negative and a positive H5N2 reference serum, together with the antigen H5N1 A/Mallard/Italy/3401/05, all purchased from the “Istituto Zooprofilattico delle Venezie”, Italy. After demonstrating the similar recognition pattern between the HA and the HACD proteins, the MAb-HA2 at a concentration of 2,5 μg/mL was selected as the capture antibody and the HRP-HA3 at a dilution of 1/20000 was selected as the detection antibody due to their optimal values of optical density at these conditions. The best dynamic range of the standard curve using the protein HACD was achieved at concentrations from 100 to 1,56 ng/mL. There were no significant differences when five batches of HACD were quantified by the ELISA sandwich and the bicinchoninic acid method linked to densitometry. In conclusion, the final parameters for the quantification of the chimeric protein HACD using an ELISA sandwich were described, which could contribute to develop a large-scale process for the final vaccine production.

Receptor–Receptor Interactions in Multiple 5-HT1A Heteroreceptor Complexes in Raphe-Hippocampal 5-HT Transmission and Their Relevance for Depression and Its Treatment. Borroto-Escuela DO, Narváez M, Ambrogini P, Ferraro L, Brito I, Romero-Fernandez W, et al. Molecules. 2018 Jun 3;23(6). pii: E1341. DOI: 10.3390/molecules23061341.

Due to the binding to a number of proteins to the receptor protomers in receptor heteromers in the brain, the term “heteroreceptor complexes” was introduced. A number of serotonin 5-HT1A heteroreceptor complexes were recently found to be linked to the ascending 5-HT pathways known to have a significant role in depression. The 5-HT1A⁻FGFR1 heteroreceptor complexes were involved in synergistically enhancing neuroplasticity in the hippocampus and in the dorsal raphe 5-HT nerve cells. The 5-HT1A protomer significantly increased FGFR1 protomer signaling in wild-type rats. Disturbances in the 5-HT1A⁻FGFR1 heteroreceptor complexes in the raphe-hippocampal 5-HT system were found in a genetic rat model of depression (Flinders sensitive line (FSL) rats). Deficits in FSL rats were observed in the ability of combined FGFR1 and 5-HT1A agonist cotreatment to produce antidepressant-like effects. It may in part reflect a failure of FGFR1 treatment to uncouple the 5-HT1A postjunctional receptors and autoreceptors from the hippocampal and dorsal raphe GIRK channels, respectively. This may result in maintained inhibition of hippocampal pyramidal nerve cell and dorsal raphe 5-HT nerve cell firing. Also, 5-HT1A⁻5-HT2A isoreceptor complexes were recently demonstrated to exist in the hippocampus and limbic cortex. They may play a role in depression through an ability of 5-HT2A protomer signaling to inhibit the 5-HT1A protomer recognition and signaling. Finally, galanin (1⁻15) was reported to enhance the antidepressant effects of fluoxetine through the putative formation of GalR1⁻GalR2⁻5-HT1A heteroreceptor complexes. Taken together, these novel 5-HT1A receptor complexes offer new targets for treatment of depression.

Recurrent Activation of Neural Circuits during Attention to Global and Local Visual Information. Iglesias-Fuster J, Piña-Novo D, Ontivero-Ortega M, Lage-Castellanos A, Valdés-Sosa M. Span J Psychol. 2018 May 28;21:E17. DOI: 10.1017/sjp.2018.9.

The attentional selection of different hierarchical level within compound (Navon) figures has been studied with event related potentials (ERPs), by controlling the ERPs obtained during attention to the global or the local echelon. These studies, using the canonical Navon figures, have produced contradictory results, with doubts regarding the scalp distribution of the effects. Moreover, the evidence about the temporal evolution of the processing of these two levels is not clear. Here, we unveiled global and local letters at distinct times, which enabled separation of their ERP responses. We combine this approach with the temporal generalization methodology, a novel multivariate technique which facilitates exploring the temporal structure of these ERPs. Opposite lateralization patterns were obtained for the selection negativities generated when attending global and local distracters (D statistics, p < .005), with maxima in right and left occipito-temporal scalp regions, respectively (η2 = .111, p < .01; η2 = .042, p < .04). However, both discrimination negativities elicited when comparing targets and distractors at the global or the local level were lateralized to the left hemisphere (η2 = .25, p < .03 and η2 = .142, p < .05 respectively). Recurrent activation patterns were found for both global and local stimuli, with scalp topographies corresponding to early preparatory stages reemerging during the attentional selection process, thus indicating recursive attentional activation. This implies that selective attention to global and local hierarchical levels recycles similar neural correlates at different time points. These neural correlates appear to be mediated by visual extra-striate areas.

Sabellastarte magnifica Carboxypetidase Inhibitor: the first Kunitz inhibitor simultaneously interacting with carboxypeptidases and serine proteases. Reytor González ML, Alonso-Del-Rivero Antigua M, Hedstrom L, Kuzmič P, Pires JR. Biochimie. 2018 May 3. pii: S0300-9084(18)30120-2. DOI: 10.1016/j.biochi.2018.04.024. [Epub ahead of print]

Multi-domain inhibitors capable to block the activity of different classes of proteases are not very common in nature. However, these kinds of molecules are attractive systems for biomedical or biotechnological applications, where two or more different targets need to be neutralized. SmCI, the Sabellastarte magnifica Carboxypeptidase Inhibitor, is a tri-domain BPTI-Kunitz inhibitor capable to inhibit serine proteases and A-like metallocarboxypeptidases. The BPTI-Kunitz family of proteins includes voltage gated channel blockers and inhibitors of serine proteases. SmCI is therefore, the only BPTI-Kunitz protein capable of inhibiting metallocarboxypeptidases. The X-ray structure of the SmCI-carboxypeptidase A complex previously obtained by us revealed that this enzyme interacts with SmCI N-tail. In the complex, the reactive loops for serine protease inhibition remain fully exposed to the solvent in each domain, suggesting SmCI can simultaneously interact with multiple serine proteases. The twofold goals of this study were: i) to establish serine proteases-SmCI binding stoichiometry, given that the inhibitor is comprised of three potential binding domains; and ii) to determine whether or not SmCI can simultaneously bind both classes of enzymes, to which it binds individually. Our experimental approach included a variety of techniques for the study of protein-protein interactions, using as model enzymes pancreatic trypsin, elastase and carboxypeptidase A. In particular, we combined information obtained from gel filtration chromatography, denaturing electrophoresis, nuclear magnetic resonance spectroscopy and enzyme inhibition assays. Our results show that SmCI is able to bind three trypsin molecules under saturating conditions, but only one elastase interacts with the inhibitor. Additionally, we demonstrated that SmCI can bind serine proteases and carboxypeptidases at the same time (at least in the ratio 1:1:1), becoming the first protease inhibitor that simultaneously block these two mechanistic classes of enzymes.

Synergistic potentiation of the anti-metastatic effect of anti EGFR mAb by its combination with immunotherapies targeting the ganglioside NGcGM3. Palomo AG, Medinilla AL, Segatori V, Barroso MDC, Blanco R, Gabri MR, et al. Oncotarget. 2018 May 8;9(35):24069–80. DOI: 10.18632/oncotarget.25290. eCollection 2018 May 8

Several Anti-EGFR mAbs are register for the treatment of human cancer. However, their impact on patients’ overall survival has been limited by tumor resistance. N-Glycolyl variant of GM3 ganglioside (NGcGM3) is specifically expressed in some human tumors, and it has been associated with a poor prognosis. Several reports have documented that GM3 physically associates to EGFR inhibiting its ligand depend phosphorylation, but it also facilitates an alternative/compensatory signaling cascade mediated by Uroquinase Plasminogen Activator Receptor (uPAR) and integrin α5β1 interaction. However, the difference between NGc and N-Acetylated (NAc) variants of GM3 regarding such interactions is unknown. We hypothesized that enrichment of NGcGM3 expression in tumors relates to advantages of this ganglioside, on ensuring both EGFR and uPAR pathways optimal function. We explored the impact of combining an anti-EGFR (7A7 mAb) with anti-NGcGM3 therapies: NGcGM3/VSSP vaccine or 14F7 mAb. Both combinations synergistically increase overall survival in two models of lung metastasis: 3LL-D122 and 4T1; but combination with NGcGM3/VSSP vaccine is significantly more effective. In 3LL-D122-metastasis, of mice treated with the best combination, both EGFR and uPAR/α5β1 integrin pathways are turn off (I.e. expression of uPAR/α5β1; and phosphorylation of EGFR, Stat3, Src and FAK are reduced); and tumor angiogenesis is decreased. Interestingly, combination treatment increases tumor infiltrating CD4⁺T, CD8⁺T and NK⁺-cells. Furthermore, a positive clinical outcome is reported for a cancer patient treated with an anti-EGFR mAb and anti-NGcGM3 therapy. Overall, our results support the combination of anti EGFR antibodies with therapies targeting NGcGM3 to increase their efficacy in future clinical trials.

System-on-chip design of the cortical-diencephalic centre of the lower urinary tract. Pérez FM, Méndez LZ, Berná Martínez JV, Lima RS. Comput Biol Med. 2018 Jun 13;99:107–122. DOI: 10.1016/j.compbiomed.2018.06.007. [Epub ahead of print]

This article presents the design of a field programmable gate array (FPGA)-based prototype of a system on chip (SoC) capable of behaving as one of the nerve centres comprising the neuroregulatory system in humans: the cortical-diencephalic nerve centre. The neuroregulatory system is a complex nerve system consisting of a heterogeneous group of nerve centres. These centres are distributed throughout the length of the spinal cord, are autonomous, communicate via interneurons, and govern and regulate the behaviour of multiple organs and systems in the human body. As a result of years of study of the functioning and composition of the neuroregulatory system of the lower urinary tract (LUT), the centres that regulate this system have been isolated. The objective of this study is to understand the individual functioning of each centre in order to create a general model of the neuroregulatory system that is capable of operating at the level of the actual nerve centre. This model represents an advancement of the current black box models that do not allow for isolated or independent treatment of system dysfunction. In this study, we re-visit our research into the viability of the hardware design of one of these centres-the cortical-diencephalic centre. We describe this hardware because functioning of the centre is completely configurable and programmable, which validates the design for other centres that comprise the neuroregulatory system. In this document, we succinctly present the formal model of the centre, propose a hardware design and an FPGA-based prototype, construct a testing and simulation environment to evaluate it and, lastly, analyse and contrast the results using data obtained from real patients, verifying that the functional behaviour fits that observed in humans.

The cytotoxic effects of propolis on breast cancer cells involve PI3K/Akt and ERK1/2 pathways, mitochondrial membrane potential, and reactive oxygen species generation. Frión-Herrera Y, Díaz-García A, Ruiz-Fuentes J, Rodríguez-Sánchez H, Sforcin JM. Inflammopharmacology. 2018 May 10. DOI: 10.1007/s10787-018-0492-y. [Epub ahead of print]

Propolis has been extensively used to improve health and prevent inflammatory diseases. Different types of Cuban propolis (red, brown and yellow) have been documented. The purpose of this research was to investigate the cytotoxic effects of Cuban red propolis (CP) on MDA MB-231 cell line, since breast cancer is considered one of the most common causes of mortality among women. Antiproliferative and cytotoxic activity of CP against MDA MB-231 cells were determined by the 3-[4,5-dimethylth-iazol-2-yl]-2,5-diphenyl tetrazoliumbromide (MTT) and lactate dehydrogenase (LDH) assays. Apoptosis/necrosis, involvement of PI3K/Akt and ERK1/2 pathways, mitochondrial membrane potential and expression of genes were investigated. CP extract exhibited antiproliferative and cytotoxic effects on MDA MB-231 cells, what may be probably related to PI3K/Akt and ERK1/2 pathways. A decreased expression of apoptosis-related genes (TP53, CASP3, BAX and P21) was seen, whereas the expressions of BCL-2, BCL-XL, NOXA and PUMA were unaffected. CP extract induced mitochondrial dysfunction and LDH release, what indicated cell necrosis associated with reactive oxygen species production and decreased cell migration. Our findings provide a basis for future investigation of chemopreventive and/or therapeutic studies against apoptosis-resistant breast cancer, in animals and humans.

The trans-[Ru(PPh₃)₂(N,N-dimethyl-N’-thiophenylthioureato-k²O,S)(bipy)]PF₆ complex has pro-apoptotic effects on triple negative breast cancer cells and presents low toxicity in vivo. Becceneri AB, Popolin CP, Plutin AM, Maistro EL, Castellano EE, Batista AA, et al. J Inorg Biochem. 2018 May 24;186:70–84. DOI: 10.1016/j.jinorgbio.2018.05.011. [Epub ahead of print]

Triple negative breast cancer (TNBC) is a heterogeneous subtype of breast tumors that does not exhibit the expression of estrogen and progesterone receptors, neither the amplification of the human epidermal growth factor receptor 2 (HER-2) gene. Despite all the advances in cancer treatments, the development of new anticancer drugs for TNBC tumors is still a challenge. There is an increasing interest in new agents to be used in cancer treatment. Ruthenium is a metal that has unique characteristics and important in vivo and in vitro results achieved for cancer treatment. Thus, in this work, with the aim to develop anticancer drugs, three new ruthenium complexes containing acylthiourea ligands have been synthesized and characterized: trans-[Ru(PPh₃)₂(N,N-dibutyl-N’-benzoylthioureato-k²O,S)(2,2′-bipyridine (bipy))]PF₆(1), trans-[Ru(PPh₃)₂(N,N-dimethyl-N’-thiophenylthioureato-k²O,S)(bipy)]PF₆(2) and trans-[Ru(PPh₃)₂(N,N-dimethyl-N’-benzoylthioureato-k²O,S)(bipy)]PF₆(3). Then, the cytotoxicity of these three new ruthenium complexes was investigated in TNBC MDA-MB-231 and in non-tumor MCF-10A cells. Complex (2) was the most selective complex and was chosen for further studies to verify its effects on cell morphology, adhesion, migration, invasion, induction of apoptosis and DNA damage in vitro, as well as its toxicity and capacity of causing DNA damage in vivo. Complex (2) inhibited proliferation, migration, invasion, adhesion, changed morphology and induced apoptosis, DNA damage and nuclear fragmentation of TNBC cells at lower concentrations compared to non-tumor MCF-10A cells, suggesting an effective action for this complex on tumor cells. Finally, complex (2) did not induce toxicity or caused DNA damage in vivo when low doses were administered to mice.

Zinc and linoleic acid pre-treatment attenuates biochemical and histological changes in the midbrain of rats with rotenone-induced Parkinsonism. Mbiydzenyuy NE, Ninsiima HI, Valladares MB, Pieme CA. BMC Neurosci. 2018 May 9;19(1):29. DOI: 10.1186/s12868-018-0429-9.

Background Studies have suggested the supplementation of Zinc and Linoleic acid in the management of neurodegenerative disorders but none has investigated the combined effects. Little is known about the neuroprotective effects of either Zinc or Linoleic acid or their combination against development of Parkinsonism. This study was designed to investigate the neuroprotective effects of Zinc and Linoleic acid in rotenone-induced Parkinsonism in rats. Methods Thirty-six young adult female rats weighing 100-150 g divided into six groups were used. Rats were induced with Parkinsonism by subcutaneous administration of rotenone (2.5 mg/kg) once a day for seven consecutive days. The rats received dimethyl sulfoxide (DMSO)/Olive oil or rotenone dissolved in DMSO/Olive oil. Groups III and IV received Zinc (30 mg/kg) or Linoleic acid (150 µl/kg) while group V received a combination of both, 2 weeks prior to rotenone injection. Groups II and VI served as negative (rotenone group) and positive (Levodopa groups) controls respectively. Oxidative stress levels were assessed by estimating Lipid peroxidation (MDA), total antioxidant capacity, Superoxide dismutase, reduced Glutathione (GSH), glutathione peroxidase and catalase in the midbrain. Histological examination was done to assess structural changes in the midbrain. Results There was a significant prevention in lipid peroxidation and decrease in the antioxidant status in intervention-treated groups as compared to the rotenone treated group. In addition, histological examination revealed that Parkinsonian rat brains exhibited neuronal damage. Cell death and reduction in neuron size induced by rotenone was prevented by treatment with zinc, linoleic acid and their combination. Conclusion These results suggest that zinc and linoleic acid and their combination showed significant neuroprotective activity most likely due to the antioxidant effect.