Fuentes et al. J Transl Med (2020) 18:414 https://doi.org/10.1186/s12967-020-02586-4 Journal of Translational Medicine PROTOCOL Open Access Glycemic variability: prognostic impact on acute ischemic stroke and the impact of corrective treatment for hyperglycemia. The GLIAS‑III translational study Blanca Fuentes1*† , Silvia Pastor‑Yborra1, Raquel Gutiérrez‑Zúñiga1, Noemí González‑Pérez de Villar2, Elena de Celis1, Jorge Rodríguez‑Pardo1, Mari Carmen Gómez‑de Frutos3, Fernando Laso‑García3, María Gutiérrez‑Fernández3, MÁngeles Ortega‑Casarrubios4, Alfonso Soto5, María López‑Fernández5, María Santamaría6, Noemí Díez‑González7, Mar M. Freijo8, Beatriz Zandio9, Raquel Delgado‑Mederos10, Ana Calleja11, Juan Carlos Portilla‑Cuenca12, Arturo Lisbona2, Laura Otero‑Ortega3*† and Exuperio Díez‑Tejedor1 Abstract Introduction: Glycemic variability (GV) represents the amplitude of oscillations in glucose levels over time and is associated with higher mortality in critically ill patients. Our aim is to evaluate the impact of GV on acute ischemic stroke (IS) outcomes in humans and explore the impact of two different insulin administration routes on GV in an animal model. Methods: This translational study consists of two studies conducted in parallel: The first study is an observational, multicenter, prospective clinical study in which 340 patients with acute IS will be subcutaneously implanted a sen‑ sor to continuously monitor blood glucose levels for 96 h. The second study is a basic experimental study using an animal model (rats) with permanent occlusion of the middle cerebral artery and induced hyperglycemia (through an intraperitoneal injection of nicotinamide and streptozotocin). The animal study will include the following 6 groups (10 animals per group): sham; hyperglycemia without IS; IS without hyperglycemia; IS and hyperglycemia without treatment; IS and hyperglycemia and intravenous insulin; and IS and hyperglycemia and subcutaneous insulin. The endpoint for the first study is mortality at 3 months, while the endpoints for the animal model study are GV, functional recovery and biomarkers. *Correspondence: blanca.fuentes@salud.madrid.org; laura.otero@salud. madrid.org † Blanca Fuentes and Laura Otero-Ortega contributed equally to this study. 1 Department of Neurology and Stroke Centre, Hospital La Paz Institute for Health Research-IdiPAZ (La Paz University Hospital, Universidad Autónoma de Madrid), Paseo de la Castellana 261, 28046 Madrid, Spain 3 Neurological Sciences and Cerebrovascular Research Laboratory, Department of Neurology and Stroke Centre, Neuroscience Area, Hospital La Paz Institute for Health Research-IdiPAZ (La Paz University Hospital, Universidad Autónoma de Madrid), Paseo de la Castellana 261, 28046 Madrid, Spain Full list of author information is available at the end of the article © The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creat​iveco​ mmons​.org/licen​ses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creat​iveco​mmons​.org/publi​cdoma​in/ zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Fuentes et al. J Transl Med (2020) 18:414 Page 2 of 8 Discussion: The GLIAS-III study will be the first translational approach analyzing the prognostic influence of GV, evaluated by the use of subcutaneous glucose monitors, in acute stroke. Trial registration https​://www.clini​caltr​ials.gov (NCT04001049) Keywords: Ischemic stroke, Glycemic variability, Insulin, Outcomes, Translational research Introduction variability. Our aim is to evaluate the impact of GV on IS Glycemic variability (GV) represents the amplitude of outcomes and examine the impact of two different insu- oscillations in blood glucose levels over time and has lin administration routes on GV in an animal model of IS. been associated with higher mortality and poor outcomes in critically ill patients [1, 2]. Few studies have evaluated Methods the prognostic influence of GV in patients with acute The translational study consists of a multicenter observa- ischemic stroke (IS) [3–8]; however, it has been suggested tional study that will include patients with acute IS and a that GV is an even more powerful prognostic factor than parallel basic experimental study using an animal model. poststroke hyperglycemia [9, 10]. Recently, the GLIAS- II study found higher GV in patients with acute IS who Design were considered nonresponders to conventional correc- Clinical study tive treatment for hyperglycemia [11]. Observational, multicenter, prospective study. However, the factors influencing the development of wider GV in acute IS remain unknown. The use of slid- Patient population The inclusion and exclusion criteria ing-scale subcutaneous insulin every 4–6 h has been sug- are listed in Table 1. gested to be associated with large oscillations in blood glucose levels and, consequently, to higher GV [12]. Simi- Randomization None (observational study). larly, intensive intravenous insulin therapy has shown higher GV than standard treatment [6] and could explain Intervention After the participants sign the informed the trend towards higher mortality in clinical trials aimed consent form, they will be implanted a CGM subcutane- at a tight reduction in blood glucose levels in acute stroke ously to monitor glucose levels for 96 h. These devices [3]. are of common use in the management of type 1 diabetes One of the limitations of previous studies that evalu- and they have been proven to be safe in stroke patients ated GV is that it was measured with intermittent read- when used in research studies. Capillary finger prick glu- ings of blood glucose levels [5, 13], with inconsistent cose levels will be recorded every 4–6 h according to the numbers of blood glucose measurements [14]. The devel- standard protocols of each center, as well as the insulin opment of continuous glucose monitors (CGM) [15, 16] administration characteristics (type, dose and route of has provided an opportunity for exploring the prognostic administration) as per the treating physician’s criteria, fol- effect of GV in acute IS and those factors that affect the lowing the local protocols for the in-hospital management oscillations in blood glucose levels during the first days of hyperglycemia. The treating physician and the nursing following a stroke. personnel will be blinded to the readings taken by the We hypothesized that GV, assessable by means of con- CMG device. All study centers will use the same brand of tinuous subcutaneous monitoring devices, could act CGM (CE-marked). All patients will be managed accord- as a powerful prognostic predictor of mortality, possi- ing to current guidelines for acute stroke management bly higher than assessment of mean or maximum blood and local protocols, including treatment with intravenous glucose levels and that the different treatment regimens thrombolysis or mechanical thrombectomy if applicable. used in routine clinical practice could modify glycemic No laboratory data other than HbA1c will be collected for Table 1 GLIAS-III main inclusion and exclusion criteria Inclusion criteria Exclusion criteria Male and female patients older than 18 years, with acute IS Transient ischemic attack Inclusion in the study within the first 24 h of stroke onset Predicted need for an MRI within the first 96 h after inclusion A prestroke score on the Modified Rankin Scale (mRS) ≤ 1 A health status, clinical condition or other characteristic that precludes appropriate Signed informed consent diagnosis, treatment, or follow-up in the trial Current drug or alcohol use dependence Participation in a therapeutic clinical trial Fuentes et al. J Transl Med (2020) 18:414 Page 3 of 8 Table 2 GLIAS-III flow diagram Baseline 96 h (± 6 h) Day 7 Day 30 (± 2 d) Day 90 or discharge (± 3 d) (± 1 d) Signed informed consent X Inclusion/exclusion criteria review X Past medical/surgical history x Pre-stroke treatments x Modified rankin scale (Pre-stroke) X X X NIHSS X X X X X Reperfusion treatment received X HbA1c X X Continuous blood glucose monitoring < –––––––––––––– > Capillary glucose levels (per local protocols) < –––––––––––––– > Insulin (type, dosage and route) administered < –––––––––––––– > according to local protocols Systemic and neurological complications < ––––––––––––––––––––––––––––––––––––––––– > Stroke etiological subtype X Secondary stroke prevention treatments X X X HbA1c glycated hemoglobin, NIHSS, National Institutes of Health Stroke Scale the study proposal. Any emergent laboratory abnormality are indexes used for studying glycemic variability in during the study follow-up will be recorded as a complica- clinical trials, and normal references values for peo- tion. ple without diabetes using CGM are available [18]. On the other hand, MAGE can provide information Study schedule The study will last 3 months per patient about the influence of the largest glycemic excur- according to the schedule shown in Table 2. sions. • Neurological or systemic complications during Primary endpoints follow-up. The following complications are pre- specified by protocol and will be checked at each • GV. Defined as the standard deviation of the mean visit: coma, seizures, early neurological impairment, blood glucose level for each patient [17]. For the main brain edema, hemorrhagic transformation, recur- analysis, we will use the GV value recorded within rent strokes, acute coronary syndrome, pulmonary the first 48 h after stroke onset. We will conduct an thromboembolism, respiratory infection, urinary exploratory analysis to evaluate the GV between 48 infection, sepsis. Any other non-pre-specified com- and 96 h. plication reported during follow-up will be also • Mortality: in-hospital and at 3 months. recorded. Secondary endpoints Data collection and management In addition to the pri- mary and secondary endpoints, the following data will be • Modified Rankin Scale (mRS) and National Institutes collected: of Health Stroke Scale. (NIHSS) at 3 months. These scales will be evaluated by a neurologist blinded to • Demographic data and risk factors: age, sex, race, the GV values. weight, height, diabetes, hypertension, dyslipidemia, • Other GV measurements: coefficient of varia- coronary arterial disease, atrial fibrillation, metabolic tion (CV), mean amplitude of glycemic excursions syndrome, chronic renal disease (eGFR of less than (MAGE); mean absolute glucose (MAG) rate of 60 ml/min/1.73m2), tobacco use and alcohol abuse. change [2, 17, 18]. CV is less influenced by mean • Pre-stroke pharmacological treatments: antiplatelets, glucose and is a good marker of hypoglycemia. A anticoagulants, blood-pressure lowering drugs, anti- threshold value to define low and high GV using CV diabetics and hypolipemiants. has recently been proposed [19]. MAGE and MAG Fuentes et al. J Transl Med (2020) 18:414 Page 4 of 8 • Stroke data: stroke severity on admission (NIHSS), Valladolid (Valladolid) and Hospital San Pedro de Alcán- date of stroke onset (or last time known asympto- tara (Cáceres). matic in those patients with unknown stroke onset), stroke etiological subtype, treatment with intrave- Experimental animal model of stroke and hyperglycemia nous thrombolysis or mechanical thrombectomy. in rats • Vital signs at baseline visit: body temperature, blood The basic experimental study will use an animal model pressure, heart rate, respiratory rate. of permanent middle cerebral artery occlusion (MCAO) and hyperglycemia in rats. The experiments are designed Data will be prospectively included in a study-specific to minimize animal suffering in compliance with our web-based database developed and managed by the Clin- Ethical Committee for the Care and Use of Animals in ical Trials Unit at Hospital 12 de Octubre which belongs Research (EU directives 86/609/CEE and 2003/65/CE). to the Spanish Clinical Research Network (SCReN), independent form the study sponsor. All data manage- Study groups A total of 60 Sprague–Dawley rats ment will follow the principles of the European regula- (8–9 weeks old, weighing 200–250 g) will be randomly tions for biomedical research ensuring confidentiality. assigned to the following groups: In compliance with European regulations/International Conference of Harmonization Good Clinical Prac- 1. Sham: hyperglycemia and surgery without MCAO. tice Guidelines, the investigator and the institution are 2. Non-stroke group: hyperglycemia without MCAO. required to permit direct access to authorized repre- 3. Normoglycemic group: normoglycemia with MCAO. sentatives of the Ethics Committee to review the subject’s 4. Non-treated hyperglycemic group: hyperglycemia original medical records for verification of study-related with MCAO. procedures and data. 5. Intravenously treated hyperglycemic group: Hyper- glycemia with MCAO and intravenous insulin Data monitoring body Monitoring will be conducted administration (dosage, 3 U/day). by dedicated personnel at La Paz University Hospital. All 6. Subcutaneously treated hyperglycemic group: Hyper- data management will follow the principles of the Euro- glycemia with MCAO and subcutaneous insulin pean regulations for biomedical research, ensuring the administration (dosage, 3 U/day). confidentiality of all personal data. In compliance with European regulations/International Conference of Har- monization Good Clinical Practice (ICHGCP) Guidelines, Randomization We will follow RIGOR recommenda- the investigator and institution are required to permit tions for animal research in terms of randomization and authorized representatives of the regulatory agency(s), blinded study [20]. Each rat will be sequentially assigned to and the IEC/IRB direct access to review the subject’s a study group based on the randomization plan. To main- original medical records for verification of study-related tain the blinding during the course of the study, we will procedures and data. ensure that the researcher who evaluates the endpoints and performs the laboratory analyses will not have access Sample size estimates Based on the data from the to the randomization codes, and a different researcher will GLIAS-II study, [11] conducted in a similar setting and perform the surgery, hyperglycemia induction and insulin using nQuery Advisor software (Statistical Solutions Ltd., administration. Following the STAIR recommendations, Cork, Ireland), we calculated that a sample of 340 patients both female and male rats will be included (1:1 ratio) [21]. will be required to achieve 80% statistical power and a The stage of estrous cycle will be recorded. If variability is confidence level alpha of 0.05 to detect a significant effect observed in the final results, a specific post-hoc analysis of of GV on mortality at 3 months. the estrous cycle stage will be performed. Recruiting hospitals Hospital Universitario La Paz Intervention Hyperglycemia will be induced by an intra- (Madrid), Hospital Universitario 12 de Octubre (Madrid), peritoneal injection of nicotinamide (210 mg/kg) (EMD Hospital Universitario A Coruña (A Coruña), Hospital Millipore, Germany). Fifteen minutes later, an intraperi- Universitario de Santiago de Compostela (Santiago de toneal injection of streptozotocin (60 mg/kg) (EMD Mil- Compostela), Hospital de Donostia (San Sebastián), Hos- lipore, Germany) will be administered [22, 23]. Blood pital Universitario Cruces (Bizkaia), Complejo Hospita- glucose levels will then be measured using a glucometer lario de Navarra (Pamplona), Hospital de la Santa Creu (ACCU-CHEK, Performa, Germany) for 72 h. The ani- i San Pau (Barcelona), Hospital Clínico Universitario de mals with a blood glucose level > 250 mg/dL will be con- sidered hyperglycemic [23]. Fuentes et al. J Transl Med (2020) 18:414 Page 5 of 8 To induce a stroke, the rats will be anesthetized via uate hindlimb functions by testing the rats’ ability to an intraperitoneal injection of a solution of ketamine traverse a wooden beam [23]. (25 mg/kg) and diazepam (2 mg/kg). Analgesia will be • Lesion size will be analyzed at 24 h and 4 weeks post- induced with a subcutaneous injection of meloxicam stroke by 7-T horizontal bore magnetic resonance (2 mg/kg). To perform the MCAO, a small craniotomy imaging (MRI) (Bruker PharmaScan, Ettlingen, Ger- will be performed, and the right middle cerebral artery many), using T2-weighted (T2-W) spin-echo ana- will be permanently ligated just before its bifurcation. tomical images, by a researcher who will be blinded Both common carotid arteries will then be occluded for to the experimental groups. 60 min, as previously described [24]. A device for CGM • Brain damage and repair-related markers will be will then be implanted subcutaneously for at least 96 h. analyzed at 4 weeks. The perilesional area of the Insulin treatment will be administered 4 h after MCAO brain will be studied in detail using immunofluores- and until 96 h later. The administration will be intrave- cence for superoxide dismutase 2, catalase, vascular nous or subcutaneous, depending on the experimental endothelial growth factor (VEGF), brain-derived group. The dosage per animal (3 U/day) has been deter- neurotrophic factor (BDNF), synaptophysin, glial mined based on the results of a previous dose–response fibrillary acidic protein, myelin basic protein and study [23]. oligodendrocyte. Immunofluorescence images will be acquired by a researcher (who will be blinded to Study schedule The study will last a total of 31 days for the experimental groups) using a Leica TCS-SPE each rat, according to the schedule illustrated in Fig. 1. confocal microscope (Leica Microsystems, Heidel- berg, Germany). Plasma levels of damage and repair Primary endpoint markers will be studied using Multiplex (eBioscience) and by ELISA (System Biosciences, Mountain View, • GV, defined as the standard deviation of the mean CA, USA). The following markers will be analyzed: glucose level [17]. We will analyze whether the insu- interleukin-1, interleukin-6, interleukin-10, vascular lin administration route (intravenous and subcutane- cell adhesion molecule, tumor necrosis factor alpha, ous) affects the GV. matrix metalloproteinase 9, VEGF, BDNF, anti-Nogo A. Secondary endpoints Sample size estimates Based on data from previous • Functional evaluation, analyzed by a researcher studies [11, 25] and using the statistical software NCSS blinded to the experimental groups, at 24 h, 1 week PASS 11 (NCSS, Utah, USA), we calculated that a sample and 4 weeks post-stroke. Motor and sensory perfor- of 10 rats per group will be required to achieve a power mance will be evaluated using a variant of Rogers’ of 80% (1-beta) and a level of significance of 5% (alpha) to functional scale [23]. The beam walking test will eval- detect a significant effect of GV on the study endpoints. Fig. 1 Experimental animal protocol. Hyperglycemia will be induced 72 h before surgery. The rats will then undergo a IS through permanent middle cerebral artery occlusion or sham surgery. Insulin treatment will be intravenously or subcutaneously administered 4 h after IS induction and up to 96 h later, after which the blood glucose levels will be continuously monitored. A functional evaluation will be performed at 24 h, 1 week and 4 weeks post-stroke. Peripheral blood will be extracted 24 h, 72 h and 4 weeks after the stroke. The MRI will be analyzed at 24 h and 4 weeks post-stroke. Four weeks after the stroke, the animals will be euthanized, and histological analyses will be performed Fuentes et al. J Transl Med (2020) 18:414 Page 6 of 8 Statistical analysis Yborra, Raquel Gutiérrez Zúñiga, Elena de Celis, María Gutiérrez Fernández, Mari Carmen Gómez de Frutos, Fernando Laso-García, Luke Diekhorst, María The statistical analysis will be performed with advice Alonso de Leciñana, Jorge Rodríguez-Pardo, Gerardo Ruiz Ares, Ricardo Rigual, from the Department of Biostatistics of La Paz Univer- Arturo Lisbona, Juan José Salazar González, Exuperio Díez-Tejedor. sity Hospital. For the continuous variables, the following Hospital Universitario 12 de Octubre: Maria Ángeles Ortega Casarrubios (Principal Investigator), Fernando Ostos-Moliz, Jaime Díaz-Guzmán, Elena information will be provided: number of participants, García Fernández, Patricia Calleja. mean, standard deviation, median, minimums, maxi- Hospital Universitario de A Coruña: Alfonso Soto (Principal Investigator), mums and 25% and 75% quartiles. For the categorical Mar Castellanos Rodrigo, María López Fernández, M Dolores Fernández Couto, Sabela Caraville Martínez, Alexia Roel García, María José Fela Painceiras, Raquel variables, the frequency distribution and 95% confidence Eiras Leal, Vanesa A Triviño-Yannuzzi, Olga Fidalgo-Baamil. interval (CI), if applicable, will be provided. The analysis Hospital Universitario de Santiago de Compostela: María Santamaría of the prognostic influence of GV on stroke outcomes (Principal Investigator), Manuel Rodríguez Yañez, Susana Arias Rivas, Emilio Rodríguez Castro, Iria López Dequit. will be performed in an exploratory manner without Hospital of Donostia: Noemí Díaz González (Principal Investigator), Maite assumptions. We will first analyze GV as a continuous Martín Zabaleta, Ana Maria de Arce Borda, Patricia de la Riva Juez, Félix variable, comparing it with primary and secondary end- González López, Miguel Urtasun Ocariz, Marta Ruibal Salgado, Marialen Arruti González. points using Student’s t-test or the Mann–Whitney U Hospital de Cruces: Mar Freijo (Principal Investigator), Alain Luna, Tomás test, as appropriate. We will also compare GV according Pérez Concha, Iratxe Ugarriza, Tirso González Pinto. to the various insulin types and administration routes Complejo Hospitalario de Navarra: Beatriz Zandio (Principal Investigator). Hospital de la Santa Creu i San Pau: Raquel Delgado Mederos (Principal applied to the patients according to the physician’s dis- Investigator), Joan Martí Fábregas, Alejandro Martínez Domeño, Pol Camps cretion. The comparisons will be performed with the Renom, Luis Prats Sánchez, Daniel Guisado Alonso, Rebeca Marín Bueno. one-way analysis of variance test. Secondly, we will per- Hospital Clínico Universitario de Valladolid: Ana Calleja (Principal Investiga‑ tor), Juan F Arenillas, Elisa Cortijo, Mercedes de Lera, Paula Simón, Javier Trigo, form a step forward logistic regression analysis for the Enrique Martínez, Alba Chavarria, Blanca Talavera, Isabel Hernández, Javier endpoints that achieved a difference with a P value < 0.1 Reyes, Beatriz Gómez. in the mean comparison tests, adjusting for age and base- Hospital San Pedro de Alcántara: Juan Carlos Portilla (Principal Investigator), Victoriano Romero Cantero, Silvia Moreno Pulido, Ignacio Casado. line National Institutes of Health Stroke Scale score. The Trial status: Recruitment started on June 30, 2020. results will be expressed as odds ratios (OR) with 95% CI. In case of lack of homogeneity in the variances, we will Authors’ contributions BF: Study concept and design, manuscript draft, project leader. SPY: study use non-parametric tests (Kruskal–Wallis and Mann– coordinator, patients’ recruitment and follow-up, revision of the manuscript Whitney test). for intellectual content. RGZ: Study concept and design, patients’ recruitment and follow-up, revision of the manuscript for intellectual content. NGP: Study concept and design, patients’ recruitment and follow-up, revision of the manuscript for intellectual content. EdC: patients’ recruitment and follow-up, DIscussion revision of the manuscript for intellectual content. Registration on the Clinical To our knowledge, few studies have used continuous glu- Trials Government website. JR: patients’ recruitment and follow-up, revision cose monitors in acute IS [7, 8, 26–28]. A relationship has of the manuscript for intellectual content. MCGF: basic experimental study (animal model), revision of the manuscript for intellectual content. FLG: basic been found between GV (by means of the standard devi- experimental study (animal model), revision of the manuscript for intellec‑ ation of mean glucose levels) and infarct growth [8], as tual content. MGF: basic experimental study (animal model), revision of the well as between GV and early neurological deterioration manuscript for intellectual content. MAOC: patients’ recruitment and follow- up, revision of the manuscript for intellectual content. AS: patients’ recruit‑ [29]. The main limitations of the prior studies using CGM ment and follow-up, revision of the manuscript for intellectual content. MLF: in acute stroke are their monocentric settings, the lack patients’ recruitment and follow-up, revision of the manuscript for intellectual of analysis of the effect of insulin administration on GV content. MS: patients’ recruitment and follow-up, revision of the manuscript for intellectual content. NDG: patients’ recruitment and follow-up, revision of and the heterogeneity in the index used to express GV, the manuscript for intellectual content. MMF: patients’ recruitment and follow- given there is no widely accepted optimal index [30]. The up, revision of the manuscript for intellectual content. BZ: patients’ recruit‑ GLIAS-III study is designed to overcome those limita- ment and follow-up, revision of the manuscript for intellectual content. RDM: patients’ recruitment and follow-up, revision of the manuscript for intellectual tions, with a prospective and multicenter approach, and content. AC: patients’ recruitment and follow-up, revision of the manuscript for is aimed at recruiting more than 300 patients. Moreover, intellectual content. JCPC: patients’ recruitment and follow-up, revision of the the development of a parallel preclinical study using an manuscript for intellectual content. AL: patients’ recruitment and follow-up, revision of the manuscript for intellectual content. LOO: Study concept and animal model of permanent MCAO will increase our design, manuscript draft, project leader for the basic experimental study in knowledge of the effect of insulin administration on animals. EDT: Study concept and design, revision of the manuscript for intel‑ GV, functional outcomes, lesion size, brain damage and lectual content. All authors read and approved the final manuscript. repair-related markers. Funding The study is investigator-initiated, promoted and coordinated by the Research Acknowledgements Foundation of La Paz University Hospital (Fundación de Investigación del We greatly appreciate the support of Morote Traducciones for editing Hospital Universitario La Paz) and funded by a grant (PI18/00991) from the assistance. Carlos III Health Institute (ISCIII) Health Care Research Fund and co-funded by GLIAS-III investigators: the European Regional Development Fund (ERDF). The study is sponsored Hospital Universitario La Paz: Blanca Fuentes (Principal Investigator), Laura by the Stroke Project of the Cerebrovascular Diseases Study Group of the Otero (Co-Principal Investigator), Noemí González Pérez de Villar, Silvia Pastor Spanish Neurological Society and the INVICTUS-Plus Spanish Network of the Fuentes et al. J Transl Med (2020) 18:414 Page 7 of 8 ISCIII (RD16/0019/0005). The study is also sponsored by Juan de la Cierva 4. Lim JS, Kim C, Oh MS, et al. Effects of glycemic variability and hyperglycemia postdoctoral fellowship (IJCI-2017-33505 to Laura Otero-Ortega, Spanish State in acute ischemic stroke on post-stroke cognitive impairments. J Diabetes Research Agency), the Spanish Ministry of Science and Innovation. Funders are Complications. 2018;32:682–7. not involved in any of the following processes: design of the trial, data collec‑ 5. Kim YS, Kim C, Jung KH, et al. Range of glucose as a glycemic variability and tion, analysis or interpretation of data neither than in writing the manuscript. 3–month outcome in diabetic patients with acute ischemic stroke. PLoS ONE. 2017;12:1–13. Availability of data and materials 6. Azevedo J, Azevedo R, Miranda M, et al. Management of hyperglycemia in After the completion of the trial, raw data will be deposited in an institutional patients with acute ischemic stroke: comparison of two strategies. Crit Care. repository and final results will be published in Open Access journals. 2009;13:P48. 7. Wada S, Yoshimura S, Inoue M, et al. Outcome prediction in acute stroke Ethics approval and consent to participate patients by continuous glucose monitoring. J Am Heart Assoc. 2018;7:1–8. The original clinical study (Protocol version 1.1; date May 6, 2019) and an 8. Shimoyama T, Kimura K, Uemura J, et al. Post stroke dysglycemia and acute amendment to include optional telematic follow-up in the pandemic COVID- infarct volume growth: a study using continuous glucose monitoring. Eur 19 scenario (Protocol version 1.2; date 24/06/2020) have been approved by Neurol. 2016;76:167–74. the Ethics Committee for Clinical Research at the La Paz University Hospital 9. Egi M, Bellomo R, Stachowski E, et al. Variability of blood glucose concen‑ and ratified by the Ethics Committees of all the participating hospitals. The tration and short-term mortality in critically ill patients. Anesthesiology. study was registered on June 27, 2019 at https​://www.clini​caltr​ials.gov (NCT 2006;105:244–52. 04001049). Recruitment started on June 30, 2020 and it is currently ongoing. 10. Mendez CE, Mok KT, Ata A, et al. Increased glycemic variability is inde‑ All the patients will be asked to sign a written informed consent form after pendently associated with length of stay andmortality in noncritically ill a detailed explanation of the nature and purpose of this study and before hospitalized patients. Diabetes Care. 2013;36:4091–7. undergoing any of the procedures related to the study. In the case of patients 11. Fuentes B, Sanz-Cuesta BE, Gutiérrez-Fernández M, et al. Glycemia in Acute with impaired consciousness, aphasia or neglect that could limit the under‑ Stroke II study: a call to improve post-stroke hyperglycemia management in standing of the study objective and procedures, the signature of the patient´s clinical practice. Eur J Neurol. 2017;24:1091–8. guardian or legal representative will be acceptable.The experimental animal 12. Hirsch IB. Sliding scale insulin—time to stop sliding. JAMA. model study was approved by the Ethics Committee for research in animals 2010;301:2009–10. at the La Paz University Hospital and authorized by the Madrid Regional 13. Yoo DS, Chang J, Kim JT, et al. Various blood glucose parameters that indi‑ Government. cate hyperglycemia after intravenous thrombolysis in acute ischemic stroke could predict worse outcome. PLoS ONE. 2014. https​://doi.org/10.1371/ Consent for publication journ​al.pone.00943​64 (Epub ahead of print 2014). Not applicable. 14. Camara-Lemarroy CR. Glucose and stroke: What about glycemic variability? J Neurol Sci. 2017;373:242–3. Competing interests 15. Finfer S, Wernerman J, Preiser J-C, et al. Clinical review: Consensus recom‑ The authors declare that they have no competing interests. mendations on measurement of blood glucose and reporting glycemic control in critically ill adults. Crit Care. 2013;17:229. Author details 16. Gomez AM, Umpierrez GE. Continuous glucose monitoring in insulin- 1 Department of Neurology and Stroke Centre, Hospital La Paz Institute treated patients in non-ICU settings. J Diabetes Sci Technol. 2014;8:930–6. for Health Research-IdiPAZ (La Paz University Hospital, Universidad Autónoma 17. Siegelaar SE, Holleman F, Hoekstra JBL, et al. Glucose variability; does it mat‑ de Madrid), Paseo de la Castellana 261, 28046 Madrid, Spain. 2 Department ter? Endocr Rev. 2010;31:171–82. of Endocrinology, Hospital La Paz Institute for Health Research-IdiPAZ (La Paz 18. Hill NR, Oliver NS, Choudhary P, et al. Normal reference range for mean University Hospital, Universidad Autónoma de Madrid), Paseo de la Castel‑ tissue glucose and glycemic variability derived from continuous glucose lana 261, 28046 Madrid, Spain. 3 Neurological Sciences and Cerebrovascular monitoring for subjects without diabetes in different ethnic groups. Diabe‑ Research Laboratory, Department of Neurology and Stroke Centre, Neu‑ tes Technol Ther. 2011;13:921–8. roscience Area, Hospital La Paz Institute for Health Research-IdiPAZ (La Paz 19. Monnier L, Colette C, Wojtusciszyn A, et al. Toward defining the threshold University Hospital, Universidad Autónoma de Madrid), Paseo de la Castellana between low and high glucose variability in diabetes. Diabetes Care. 261, 28046 Madrid, Spain. 4 Department of Neurology, Hospital Universitario 2017;40:832–8. 12 de Octubre, Universidad Complutense de Madrid, Madrid, Spain. 5 Depart‑ 20. Lapchak PA, Zhang JH, Noble-Haeusslein LJ. RIGOR guidelines: escalat‑ ment of Endocrinology, Hospital Universitario A Coruña, A Coruña, Spain. ing STAIR and STEPS for effective translational research. Transl Stroke Res. 6 Department of Neurology, Hospital La Paz Institute for Health Research- 2013;4:279–85. IdiPAZ (La Paz University Hospital, Universidad Autónoma de Madrid), Santiago 21. Fisher M, Feuerstein G, Howells DW, et al. Update of the stroke therapy de Compostela, Spain. 7 Department of Neurology, Hospital de Donostia, San academic industry roundtable preclinical recommendations. Stroke. Sebastián, Spain. 8 Biocruces Bizkaia Health Research Institute, Department 2009;40:2244–50. of Neurology, Hospital Universitario Cruces, Bizkaia, Spain. 9 Department 22. Szkudelski T. Streptozotocin-nicotinamide-induced diabetes in the rat. of Neurology, Complejo Hospitalario de Navarra, Pamplona, Spain. 10 Depart‑ Characteristics of the experimental model. Exp Biol Med. 2012;237:481–90. ment of Neurology, Hospital de la Santa Creu i San Pau, Barcelona, Spain. 23. Qinna NA, Badwan AA. Impact of streptozotocin on altering normal glucose 11 Department of Neurology, Hospital Clínico Universitario de Valladolid, Val‑ homeostasis during insulin testing in diabetic rats compared to normogly‑ ladolid, Spain. 12 Department of Neurology, Hospital San Pedro de Alcántara, cemic rats. Drug Des Devel Ther. 2015;9:2515–25. Cáceres, Spain. 24. Gutiérrez-Fernández M, Rodríguez-Frutos B, Alvarez-Grech J, et al. Functional recovery after hematic administration of allogenic mesenchymal stem cells Received: 24 August 2020 Accepted: 24 October 2020 in acute ischemic stroke in rats. Neuroscience. 2011;175:394–405. 25. Gómez-De Frutos MC, Laso-Garciá F, Diekhorst L, et al. Intravenous delivery of adipose tissue-derived mesenchymal stem cells improves brain repair in hyperglycemic stroke rats. Stem Cell Res Ther. 2019;10:1–13. 26. Allport L, Baird T, Butcher K, et al. Frequency and temporal profile of post‑ References stroke hyperglycemia using continuous glucose monitoring. Diabetes Care. 1. Krinsley JS. Glycemic variability: a strong independent predictor of mortality 2006;29:1839–44. in critically ill patients. Crit Care Med. 2008;36:3008–13. 27. Ribo M, Molina CA, Delgado P, et al. Hyperglycemia during ischemia rapidly 2. Hermanides J, Vriesendorp TM, Bosman RJ, et al. Glucose variability is associ‑ accelerates brain damage in stroke patients treated with tPA. J Cereb Blood ated with intensive care unit mortality*. Crit Care Med. 2010;38:838–42. Flow Metab. 2007;27:1616–22. 3. González-Moreno EI, Cámara-Lemarroy CR, González-González JG, et al. Gly‑ 28. Baird TA, Parsons MW, Phanh T, et al. Persistent poststroke hyperglycemia cemic variability and acute ischemic stroke: the missing link? Transl Stroke is independently associated with infarct expansion and worse clinical Res. 2014;5:638–46. outcome. Stroke. 2003;34:2208–14. Fuentes et al. J Transl Med (2020) 18:414 Page 8 of 8 29. Hui J, Zhang J, Mao X, et al. The initial glycemic variability is associated with Publisher’s Note early neurological deterioration in diabetic patients with acute ischemic Springer Nature remains neutral with regard to jurisdictional claims in pub‑ stroke. Neurol Sci. 2018;39:1571–7. lished maps and institutional affiliations. 30. Palaiodimou L, Lioutas VA, Lambadiari V, et al. Glycemia management in acute ischemic stroke: current concepts and novel therapeutic targets. Postgrad Med. 2019;131:423–37. Ready to submit your research ? Choose BMC and benefit from: • fast, convenient online submission • thorough peer review by experienced researchers in your field • rapid publication on acceptance • support for research data, including large and complex data types • gold Open Access which fosters wider collaboration and increased citations • maximum visibility for your research: over 100M website views per year At BMC, research is always in progress. Learn more biomedcentral.com/submissions