title: Leg ischaemia before circulatory arrest alters brain leucocyte count and respiratory chain redox state.
abstract: OBJECTIVE
  Remote ischaemic preconditioning and its neuroprotective abilities are currently under investigation and the method has shown significant effects in several small and large animal studies. In our previous studies, leucocyte filtration during cardiopulmonary bypass reduced cerebrocortical adherent leucocyte count and mitigated cerebral damage after hypothermic circulatory arrest (HCA) in piglets. This study aimed to obtain and assess direct visual data of leucocyte behaviour in cerebral vessels after hypothermic circulatory arrest following remote ischaemic preconditioning.
  
  
  METHODS
  Twelve native stock piglets were randomized into a remote ischaemic preconditioning group (n = 6) and a control group (n = 6). The intervention group underwent hind-leg ischaemia, whereas the control group received a sham-treatment before a 60-min period of hypothermic circulatory arrest. An intravital microscope was used to obtain measurements from the cerebrocortical vessel in vivo. It included three sets of filters: a violet filter to visualize microvascular perfusion and vessel diameter, a green filter for visualization of rhodamine-labelled leucocytes and an ultraviolet filter for reduced nicotinamide adenine dinucleotide (NADH) analysis. The final magnification on the microscope was 400. After the experiment, cerebral and cerebellar biopsies were collected and analysed with transmission electron microscope by a blinded analyst.
  
  
  RESULTS
  In the transmission electron microscope analysis, the entire intervention group had normal, unaffected rough endoplasmic reticulum's in their cerebellar tissue, whereas the control group had a mean score of 1.06 (standard deviation 0.41) (P = 0.026). The measured amount of adherent leucocytes was lower in the remote ischaemic preconditioning group. The difference was statistically significant at 5, 15 and 45 min after circulatory arrest. Statistically significant differences were seen also in the recovery phase at 90 and 120 min after reperfusion. Nicotinamide adenine dinucleotide autofluorescence had statistically significant differences at 10 min after cooling and at 120 and 180 min after hypothermic circulatory arrest.
  
  
  CONCLUSIONS
  Remote ischaemic preconditioning seems to provide better mitochondrial respiratory chain function as indicated by the higher NADH content. It simultaneously provides a reduction of adherent leucocytes in cerebral vessels after hypothermic circulatory arrest. Additionally, it might provide some degree of cellular organ preservation as implied by the electron microscopy results.
authors.affiliation: Department of Surgery, Oulu University Hospital, Oulu, Finland.
authors.forename: Fredrik S
authors.initials: FS
authors.lastname: Yannopoulos
authors.forename: Oiva
authors.initials: O
authors.lastname: Arvola
authors.forename: Henri
authors.initials: H
authors.lastname: Haapanen
authors.forename: Johanna
authors.initials: J
authors.lastname: Herajärvi
authors.forename: Ilkka
authors.initials: I
authors.lastname: Miinalainen
authors.forename: Hanna
authors.initials: H
authors.lastname: Jensen
authors.forename: Kai
authors.initials: K
authors.lastname: Kiviluoma
authors.forename: Tatu
authors.initials: T
authors.lastname: Juvonen
mesh_terms: D000818:Animals; D001921:Brain / Q000098:blood supply* / Q000378:metabolism / Q000648:ultrastructure; D002545:Brain Ischemia / Q000209:etiology / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D002448:Cell Adhesion; D002560:Cerebrovascular Circulation; D004195:Disease Models, Animal; D004579:Electron Transport; D005260:Female; D006324:Heart Arrest, Induced; D007035:Hypothermia; D019194:Ischemic Preconditioning / Q000379:methods*; D007958:Leukocyte Count; D036904:Leukocyte Rolling; D007962:Leukocytes; D035002:Lower Extremity / Q000098:blood supply*; D008833:Microcirculation; D046529:Microscopy, Electron, Transmission; D008928:Mitochondria / Q000378:metabolism*; D010084:Oxidation-Reduction; D012039:Regional Blood Flow; D015427:Reperfusion Injury / Q000209:etiology / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D013552:Swine; D013997:Time Factors
keywords: Hypothermia/circulatory arrest; Ischaemia/reperfusion injury brain; Preconditioning brain