Computerised image analysis in conjunction with fluorescence microscopy for the study of blood-brain barrier permeability in vivo

The present paper describes a new method using computerised image analysis techniques for quantification of tracer extravasation over the blood-brain barrier as studied by intravital fluorescence microscopy. Cats were equipped with an open cranial window and continuously infused with fluorescein isothiocyanate-labelled dextran (FITC-dextran, mol. wt. 70 000) to maintain a steady plasma concentration. Several cortical fields were recorded in each experiment and the images stored on video tape for off-line analysis. This procedure, which largely eliminates the superficial pial vasculature and allows extraction of the extravasation areas, consists of the following steps: (1) averaging of images, (2) software shading correction based on the original images for compensation of optical non-uniformity, (3) correction of displacement artefacts, (4) intensity adjustment, (5) generation of subtraction images by subtracting the first image of a series from the subsequent ones, (6) median filtering and thresholding, (7) a length recognition algorithm, and (8) elimination of small areas. Compared to the previously described method, step (2) has been newly developed and steps (4) and (8) added to enhance sensitivity for detecting tracer extravasation. The degree of extravasation in a cortical field at a given time point [E(f) value] was calculated as the mean intensity of the remaining pixels. TheE(f) is a quantitative value computed by a fully automatised procedure which takes into account the number, as well as the size and intensity, of extravasation areas in a given cortical field. TheE(f) values obtained at different times in a series of experiments were averaged to give theE(I) value. TheE(I) value did not alter when hypercapnia was employed to induce pure vasodilatation. On the other hand it increased dramatically, indicating tracer extravasation, during topical application of high concentrations of adenosine (10^–5–10^–3 M). The new computerised image analysis procedure may therefore be suitable for measuring quantitatively tracer extravasation over the blood-brain barrier in vivo under different experimental conditions. It may also be applicable to study changes of vascular permeability in peripheral vascular beds.     

Thermal and respiratory control in young rats with altered caloric intake during postnatal development

We asked to what extent differences in caloric intake during the first postnatal weeks may modify thermal and respiratory control of 1-month old rats. Large-size (Large) and small-size (Small) rats were obtained by raising rats in, respectively, small (6 pups) and large (16 pups) litters. In Small, the rate of oxygen consumption (V(O(2))/kg) was less than in Large during the first 2-3 weeks, and higher thereafter, when the thermogenic needs to maintain body temperature (Tb) increased. At day 31, when body weight in Small was approximately 80% of Large, Small maintained Tb in the cold with higher V(O(2))/kg than Large. The total uncoupling protein of the brown adipose tissue was unchanged. Also pulmonary ventilation (VE/kg) was higher in Small, maintaining the proportionality with V(O(2)). Lung weight in Small was reduced in proportion to body weight, with higher protein-DNA ratio. The compliances of the respiratory system and lungs, normalized by body weight, and the hyperventilatory responses to hypoxia or hypercapnia, expressed as % increase in VE/V(O(2)), were similar in Small and Large. Differences between Small and Large were reduced or no longer present in a group of Small rats raised until their body weight was as in Large. We conclude that rather important developmental differences in caloric intake and metabolic level, in otherwise healthy rats, had no long-term carry over effects in the developmental processes of respiratory and thermal control, other than the effects strictly attributable to the alterations in body size.     

Elimination of CO2 insufflation-induced hypercapnia in open heart surgery using an additional venous reservoir

 Open in a separate windowOBJECTIVESCarbon dioxide (CO₂) gas insufflation is used for continuous de-airing during open heart surgery. The aim was to evaluate if an additional separate venous reservoir eliminates CO₂ insufflation-induced hypercapnia and keeps sweep gas flow of the oxygenator constant.METHODSA separate reservoir was used during cardiopulmonary bypass in addition to a standard venous reservoir. The additional reservoir received drained blood and CO₂ gas continuously via a suction drain (1 l/min) and handheld suction devices from the surgical wound. CO₂ gas was insufflated via a gas diffuser in the open wound at 10 l/min. In a cross-over design for each patient, gas and blood were either continuously drained from the additional to the standard venous reservoir or not. CO₂ pressure in arterial blood (PaCO₂) was measured after adjustment of sweep gas flow as necessary and after steady state of PaCO₂ was observed. Mean values for each setup (median 4 times) for each patient were analysed with Wilcoxon rank-sum test.RESULTSTen adult patients undergoing open aortic valve replacement were included. Median PaCO₂ did not differ between setups (5.41; 5.29–5.57, interquartile range vs 5.41; 5.24–5.58, P = 0.92), whereas sweep gas flow (l/min) was lower (2.58; 2.50–3.16 vs 4.42; 4.0–5.40, P = 0.002) when CO₂ gas was not drained from the additional to the standard reservoir.CONCLUSIONSAn additional venous reservoir for the evacuation of blood from the open surgical wound eliminates CO₂ insufflation-induced hypercapnia in open heart surgery keeping PaCO₂ and sweep gas flow constant. This prevents possible CO₂-induced hyperperfusion of the brain and decreases the risk of cerebral particulate embolization during CO₂ insufflation for de-airing in open heart surgery.Clinical trial registration{"type":"clinical-trial","attrs":{"text":"NCT04202575","term_id":"NCT04202575"}}NCT04202575.IRB approval dat and number2018-07-13 and 2018/1091-31.     

无创正压通气治疗慢性阻塞性肺疾病高碳酸血症的疗效观察

目的探讨无创正压通气治疗慢性阻塞性肺疾病（COPD）高碳酸血症的价值.方法58例COPD二氧化碳潴留患者随机分入治疗组及对照组,对照组仅行常规药物治疗,治疗组在常规药物治疗的基础上加用无创正压通气治疗.比较治疗开始2小时、24小时、7天两组同期血气、血压、心率、呼吸频率等改善情况.结果治疗组同期血气、血压、心率、呼吸频率等较对照组明显改善（P均＜0.05）.结论应用无创通气可明显改善COPD患者症状,提高PO2降低PCO2,对COPD高碳酸血症患者有确切疗效.     

无创正压通气救治慢性阻塞性肺疾病呼吸衰竭高碳酸血症昏迷的疗效观察

目的观察无创正压通气（NPPV）治疗慢性阻塞性肺疾病（COPD）呼吸衰竭昏迷患者的疗效。方法对入选的32例COPDⅡ型呼吸衰竭昏迷患者和42例COPDⅡ型呼吸衰竭非昏迷患者在常规治疗的基础上行双水平无创正压辅助通气（BiPAP），动态观察治疗前后动脉血气、Glasgow昏迷评分（GCS）、NPPV天数、有效率及不良反应情况。结果昏迷组32例患者中30例经BiPAP通气后好转（2h后意识转清26例，3．8h意识转清3例，24h后完全清醒1例），血气分析指标好转，潮气量、每分通气量增加，通气（9±4）d恢复至稳定期状态，与非昏迷组（7±3）d比较差异无统计学意义（P〉0．05）。但昏迷组胃肠胀气的发生率高，较非昏迷组明显增加（80．5％比10．6％，P〉0．05）。两组间成功率没有明显差异（93．75％比97．62％，P〉0．05）。结论昏迷不是NPPV治疗的绝对禁忌证。     

Hypoxic tolerance in air-breathing invertebrates

Terrestrial invertebrates experience hypoxia in many habitats and under a variety of physiological conditions. Some groups (at least insects) are much more capable of recovery from anoxia than most vertebrates, but there is still a tremendous unexplained variation in hypoxia/anoxia tolerance among terrestrial invertebrates. Crustaceans and arachnids may be less often confronted with hypoxic environments than insects and myriapods and also seem to be less hypoxia/anoxia tolerant. Tracheated groups, especially those that are able to ventilate their tracheal system like many insects, cope with lower critical PO2 than nontracheated groups. Modulation of oxygen carrier proteins is normally not important in hypoxia resistance. Recent application of genetic and cellular tools are revealing that many of the same pathways documented for mammals (e.g. HIF, nitric oxide) function to regulate morphological and biochemical responses to hypoxia/anoxia in insects.     

无创正压通气治疗拒绝插管的COPD急性加重并重度高碳酸血症呼吸衰竭患者的临床观察

目的探讨应用无创正压通气(NiPPV)治疗慢性阻塞性肺疾病急性加重(AECOPD)并重度高碳酸血症呼吸衰竭(HRF)的可行性。方法选取AECOPD并重度HRF患者12例,常规治疗时选其中6例(通气组)应用中等吸氧浓度(FiO2)与NiPPV治疗,注意管理气道(引流排痰、扩张气道)、管理面罩(防漏气)、防治通气并发症,另外6例为对照组。结果通气组6例患者中3例在通气治疗2h后动脉血气好转,3例于12h后好转,神志转清,36h后血气稳定,48～72h后,5例逐渐成功脱机最后好转出院,1例在第7天死亡。对照组中2例于治疗36h后血气好转,第5天出院,4例死亡。结论应用NiPPV可以成功抢救AECOPD并重度HRF患者,其难点在于排痰通畅气道,同时应用中等FiO2改善缺氧意义重大,其成功率取决于包括NiPPV在内的综合疗效。     

高碳酸血症对急性肺损伤保护作用及机制探讨的实验研究

目的探讨高碳酸血症对急性肺损伤模型的保护作用及可能的机制。方法24只新西兰兔按随机数字表法分为对照组、治疗组、预防组，每组8只。采用脂多糖静脉注射复制肺损伤模型，观察3组兔血流动力学、血气指标的变化；检测肺组织湿／干重（W／D）比、光镜、肺损伤组织学定量评价指标（IQA）来评估肺脏的损伤程度。通过对肺组织中髓过氧化物酶（MPO）、丙二醛（MDA）及血清和支气管肺泡灌洗液（BALF）中白细胞介素-8（IL-8）、肿瘤坏死因子-α（TNF—α）浓度及中性粒细胞凋亡率的测定，阐明高碳酸血症对肺损伤保护的可能机制。结果（1）治疗组、预防组在模型形成后平均动脉压、心率、动脉血二氧化碳分压（PaCO2）、动脉血氧分压／吸入气氧浓度（PaO2／FiO2）分别为（79±6）mm Hg（1mmHg=0．133kPa）、（180±10）次／min、（99±13）mmHg、250±26，（80±9）mmHg、（181±12）次／min、（95±11）mmHg、241±56，与对照组[（66±10）mmHg、（139±13）次／min、（31±4）mmHg、182±35]比较差异有统计学意义（t值分别为4．05、26．32、5．36、28．15、12．54、11．07、16．13、12．36。P均分别〈0．05、0．01）；（2）治疗组、预防组W／D、NPO、MDA分别为1．98±0．28、1．87±0．30、（6．1±1．6）U／g、（5．8±1．5）U／g、（20±5）mg／L、（19±4）mg／L，与对照组[2．43±0．26、（9．0±1．3）U／g、（36±8）mg／L]比较差异有统计学意义（t值分别为11．07、24．46、2．35、9．63，12．34、25．32，P分别〈0．05、0．01）；（3）治疗组、预防组血清和BALF中IL-8、TNF—α、中性粒细胞凋亡率分别为（50±8）ng／ml、（103±49）ng／ml、（94±16）ng／ml、（44±9）ng／ml、（38±9）％、（56±5）％、（49±7）ng／ml、（96±50）ng／ml、（91±14）ng／ml、（39±6）ng／ml、（39±10）％、（55±10）％，与对照组[（91±43）ng／ml、（177±60）ng／ml、（162±15）ng／ml、（67±7）ng／ml、（19±7）％、（43±7）％]比较差异有统计学意义（t值分别为7．12、5．55、7．30、3．93、13．08、8．00，P分别〈0．05、0．01）；（4）各组血清及BALF中IL-8、TNF—α与中性粒细胞凋亡率呈负相关（r值分别为-0．73、-0．72、-0．52、-0．64、-0．73、-0．56、-0．57、-0．78、-0．69、-0．75、-0．82、-0．84，P均〈0．05）。结论高碳酸血症对急性肺损伤具有保护作用，对血流动力学无明显影响。