Morphologic Mapping of the Sublingual Microcirculation in Healthy Volunteers

PURPOSE Monitoring the sublingual and oral microcirculation (SM-OM) using hand-held vital microscopes (HVMs) has provided valuable insight into the (patho)physiology of diseases. However, the microvascular anatomy in a healthy population has not been adequately described yet. METHODS Incident dark field-based HVM imaging was used to visualize the SM-OM. First, the SM was divided into four different fields; Field-a (between incisors-lingua), Field-b (between the canine-first premolar-lingua), Field-c (between the first-second premolar-lingua), Field-d (between the second molar-wisdom teeth-lingua). Second, we investigated the buccal area, lower and upper lip. Total/functional vessel density (TVD/FCD), focus depth (FD), small vessel mean diameters (SVMDs), and capillary tortuosity score (CTS) were compared between the areas. RESULTS Fifteen volunteers with a mean age of 29 ± 6 years were enrolled. No statistical difference was found between the sublingual fields in terms of TVD (p = 0.30), FCD (p = 0.38), and FD (p = 0.09). SVMD was similar in Field-a, Field-b, and Field-c (p = 0.20-0.30), and larger in Field-d (p < 0.01, p = 0.015). The CTS of the buccal area was higher than in the lips. CONCLUSION The sublingual area has a homogenous distribution in TVD, FCD, FD, and SVMD. This study can be a description of the normal microvascular anatomy for future researches regarding microcirculatory assessment

Identifying a sublingual triangle as the ideal site for assessment of sublingual microcirculation

The sublingual mucosa is a commonly used intraoral location for identifying microcirculatory alterations using handheld vital microscopes (HVMs). The anatomic description of the sublingual cave and its related training have not been adequately introduced. The aim of this study was to introduce anatomy guided sublingual microcirculatory assessment. Measurements were acquired from the floor of the mouth using incident dark-field (IDF) imaging before (T0) and after (T1) sublingual cave anatomy instructed training. Instructions consists of examining a specific region of interested identified through observable anatomical structures adjacent and bilaterally to the lingual frenulum which is next to the sublingual papilla. The anatomical location called the sublingual triangle, was identified as stationed between the lingual frenulum, the sublingual fold and ventrally to the tongue. Small, large, and total vessel density datasets (SVD, LVD and TVD respectively) obtained by non-instructed and instructed measurements (NIN (T0) and IM (T1) respectively) were compared. Microvascular structures were analyzed, and the presence of salivary duct-related microcirculation was identified. A total of 72 video clips were used for analysis in which TVD, but not LVD and SVD, was higher in IM compared to NIM (NIM vs. IM, 25 ± 2 vs. 27 ± 3 mm/mm$^{2}$ (p = 0.044), LVD NIM vs. IM: 7 ± 1 vs. 8 ± 1mm/mm$^{2}$ (p = 0.092), SVD NIM vs. IM: 18 ± 2 vs. 20 ± 3 mm/mm$^{2}$ (p = 0.103)). IM resulted in microcirculatory assessments which included morphological properties such as capillaries, venules and arterioles, without salivary duct-associated microcirculation. The sublingual triangle identified in this study showed consistent network-based microcirculation, without interference from microcirculation associated with specialized anatomic structures. These findings suggest that the sublingual triangle, an anatomy guided location, yielded sublingual based measurements that conforms with international guidelines. IM showed higher TVD values, and future studies are needed with larger sample sizes to prove differences in microcirculatory parameters

Microcirculatory alterations in critically ill COVID-19 patients analyzed using artificial intelligence

Background: The sublingual microcirculation presumably exhibits disease-specific changes in function and morphology. Algorithm-based quantification of functional microcirculatory hemodynamic variables in handheld vital microscopy (HVM) has recently allowed identification of hemodynamic alterations in the microcirculation associated with COVID-19. In the present study we hypothesized that supervised deep machine learning could be used to identify previously unknown microcirculatory alterations, and combination with algorithmically quantified functional variables increases the model's performance to differentiate critically ill COVID-19 patients from healthy volunteers. Methods: Four international, multi-central cohorts of critically ill COVID-19 patients and healthy volunteers (n = 59/n = 40) were used for neuronal network training and internal validation, alongside quantification of functional microcirculatory hemodynamic variables. Independent verification of the models was performed in a second cohort (n = 25/n = 33). Results: Six thousand ninety-two image sequences in 157 individuals were included. Bootstrapped internal validation yielded AUROC(CI) for detection of COVID-19 status of 0.75 (0.69-0.79), 0.74 (0.69-0.79) and 0.84 (0.80-0.89) for the algorithm-based, deep learning-based and combined models. Individual model performance in external validation was 0.73 (0.71-0.76) and 0.61 (0.58-0.63). Combined neuronal network and algorithm-based identification yielded the highest externally validated AUROC of 0.75 (0.73-0.78) (P < 0.0001 versus internal validation and individual models). Conclusions: We successfully trained a deep learning-based model to differentiate critically ill COVID-19 patients from heathy volunteers in sublingual HVM image sequences. Internally validated, deep learning was superior to the algorithmic approach. However, combining the deep learning method with an algorithm-based approach to quantify the functional state of the microcirculation markedly increased the sensitivity and specificity as compared to either approach alone, and enabled successful external validation of the identification of the presence of microcirculatory alterations associated with COVID-19 status. Keywords: Artificial intelligence; COVID-19; Deep learning; Microcirculation; Neuronal network

Intraoperative Imaging Techniques to Visualize Hepatic (Micro)Perfusion: An Overview

The microcirculation plays a crucial role in the distribution of perfusion to organs. Studies have shown that microcirculatory dysfunction is an independent predictor of morbidity and mortality. Hence, ass

Hemodilution causes glycocalyx shedding without affecting vascular endothelial barrier permeability in rats

Background The consequences of acute normovolemic hemodilution (ANH) following different types of fluids on the different components of the glycocalyx and on vascular barrier permeability (VBP) remain unknown. Aim The aim of the study was to investigate whether the microcirculatory disruption and glycocalyx shedding induced by ANH alters VBP and whether this is affected by the composition and volume of the resuscitation fluid. Materials and Methods Anesthetized Wistar albino rats (n=24) underwent stepwise ANH at hematocrit levels of 35%, 25%, 20%, and 15% induced by the exchange of blood with 6% balanced hydroxyethyl starch (1:1), balanced crystalloid (1:3), and normal saline (NS) (1:3). Glycocalyx-shed products were measured at each level of hemodilution. VBP was reflected in the decay of fluorescence dyes of different molecular size and their plasma retention ratios. Edema was assessed by measuring organ water content and muscle microcirculation by hand-held videomicroscopy. Results NS caused increased degradation of heparan sulfate and hyaluronan compared with the control group (P=0.003, P=0.004, respectively). Neither VBP nor tissue edema was affected by the fluid used. The total and perfused vessel densities within the microcirculation of muscle tissue decreased at hematocrit 15% in the balanced crystalloid (P=0.02) and NS groups only (P<0.0001, P=0.0003, respectively) compared with baseline. Conclusions Balanced colloid solution preserved the glycocalyx layer better than balanced and unbalanced crystalloid solutions while maintaining the microcirculatory function associated with an improved total intravascular volume. Among the fluids tested, NS caused the most microcirculatory alterations. While ANH caused the degradation of glycocalyx components regardless of fluid, it did not disrupt the vascular barrier as indicated by macromolecular leakage. Relevance for Patients The results of this study provide insight into the choice of fluid for optimal perioperative fluid management and the consequences of fluid type on the vascular barrier, glycocalyx, and microcirculation

Real-time observation of microcirculatory leukocytes in patients undergoing major liver resection

Ischemia/reperfusion injury and inflammation are associated with microcirculatory dysfunction, endothelial injury and glycocalyx degradation. This study aimed to assess microcirculation in the sublingual, intestinal and the (remnant) liver in patients undergoing major liver resection, to define microcirculatory leukocyte activation and its association with glycocalyx degradation. In this prospective observational study, the microcirculation was assessed at the beginning of surgery (T0), end of surgery (T1) and 24 h after surgery (T2) using Incident Dark Field imaging. Changes in vessel density, blood flow and leukocyte behaviour were monitored, as well as clinical parameters. Syndecan-1 levels as a parameter of glycocalyx degradation were analysed. 19 patients were included. Sublingual microcirculation showed a significant increase in the number of rolling leukocytes between T0 and T1 (1.5 [0.7–1.8] vs. 3.7 [1.7–5.4] Ls/C-PCV/4 s respectively, p = 0.001), and remained high at T2 when compared to T0 (3.8 [3–8.5] Ls/C-PCV/4 s, p = 0.006). The microvascular flow decreased at T2 (2.4 ± 0.3 vs. baseline 2.8 ± 0.2, respectively, p < 0.01). Duration of vascular inflow occlusion was associated with significantly higher numbers of sublingual microcirculatory rolling leukocytes. Syndecan-1 increased from T0 to T1 (42 [25–56] vs. 107 [86–164] ng/mL, p < 0.001). The microcirculatory perfusion was characterized by low convection capacity and high number of rolling leukocytes. The ability to sublingually monitor the rolling behaviour of the microcirculatory leukocytes allows for early identification of patients at risk of increased inflammatory response following major liver resection

Interpatient heterogeneity in hepatic microvascular blood flow during vascular inflow occlusion (Pringle manoeuvre)

Background: Vascular inflow occlusion (VIO) during liver resections (Pringle manoeuvre) can be applied to reduce blood loss, however may at the same time, give rise to ischemia-reperfusion injury (IRI). The aim of this study was to assess the characteristics of hepatic microvascular perfusion during VIO in patients undergoing major liver resection. Methods: Assessment of hepatic microcirculation was performed using a handheld vital microscope (HVM) at the beginning of surgery, end of VIO (20 minutes) and during reperfusion after the termination of VIO. The microcirculatory parameters assessed were: functional capillary density (FCD), microvascular flow index (MFI) and sinusoidal diameter (SinD). Results: A total of 15 patients underwent VIO; 8 patients showed hepatic microvascular perfusion despite VIO (partial responders) and 7 patients showed complete cessation of hepatic microvascular perfusion (full responders). Functional microvascular parameters and blood flow levels were significantly higher in the partial responders when compared to the full responders during VIO (FCD: 0.84±0.88 vs. 0.00±0.00 mm/mm2, P<0.03, respectively, and MFI: 0.69-0.22 vs. 0.00±0.00, P<0.01, respectively). Conclusions: An interpatient heterogeneous response in hepatic microvascular blood flow was observed upon VIO. This may explain why clinical strategies to protect the liver against IRI lacked consistency

Recruitment of sublingual microcirculation using handheld incident dark field imaging as a routine measurement tool during the postoperative de-escalation phase—a pilot study in post ICU cardiac surgery patients

Abstract Background Management of tissue perfusion following cardiac surgery is a challenging task where common clinical parameters do not reflect microcirculatory dysfunction. Heterogeneity in blood flow perfusion and abnormalities in capillary density characterize microcirculatory dysfunction. The restoration of a normal microcirculation may become a novel target for therapy in the future in addition to macrocirculatory parameters. The aim of this study is to determine how the sublingual microcirculatory parameters vary at the bedside in post-cardiac surgery patients which underwent diuretic therapy to correct fluid overload. Methods In this prospective observational pilot study, video clips of sublingual microcirculation in post-cardiac surgery patients receiving furosemide and/or spironolactone to achieve normal fluid balance were recorded using Cytocam-IDF imaging. Data was obtained on the first (T0), second (T1), and third (T2) day after the patients left the intensive care unit (ICU). Measurements were analyzed off-line to obtain the following microcirculatory parameters: total vessel density (TVD), microcirculatory flow index (MFI), proportion of perfused vessel (PPV), and perfused vessel density (PVD). Macrocirculatory parameters and body weight were also collected at these time points. Results Ninety measurements were performed in ten post ICU cardiac surgery patients. Thirteen measurements were excluded due to quality reasons; these excluded measurements were spread across the patients and time points, and there was no loss of patients or time points. An increase in TVD was observed from T0 to T1 (20 ± 2.7 to 24 ± 3.2 mm/mm2; p = 0.0410) and from T0 to T2 (20 ± 2.7 to 26 ± 3.3 mm/mm2; p = 0.0005). An increase in PVD was present from T0 to T1 (19 ± 2.3 to 24 ± 3.5 mm/mm2; p = 0.0072) and from T0 to T2 (19 ± 2.3 to 26 ± 3.4 mm/mm2, p = 0.0008). Fluid overload was assessed through a positive cumulative fluid balance on the day of ICU discharge. Conclusions Cytocam-IDF imaging to monitor microcirculation as a daily parameter is feasible and could become a valuable tool to non-invasively assess the tissue oxygenation at the bedside. An increase in TVD and PVD (functional capillary density) indicated the recruitment of the sublingual microcirculation in patients with diuretic therapy. Future research is needed to prove the correlation between the recruitment of the sublingual microcirculation and the de-escalation phase of the fluid management

Leukocyte-Endothelium Interaction in the Sublingual Microcirculation of Coronary Artery Bypass Grafting Patients

markdownabstractObjective: The aim of this study was to apply an innovative methodology to incident dark-field (IDF) imaging in coronary artery bypass grafting (CABG) patients for the identification and quantification of rolling leukocytes along the sublingual microcirculatory endothelium. Methods: This study was a post hoc analysis of a prospective study that evaluated the perioperative course of the sublingual microcirculation in CABG patients. Video images were captured using IDF imaging following the induction of anesthesia (T0) and cardiopulmonary bypass (CPB) (T1) in 10 patients. Rolling leukocytes were identified and quantified using frame averaging, which is a technique that was developed for correctly identifying leukocytes. Results: The number of rolling leukocytes increased significantly from T0 (7.5 [6.4-9.1] leukocytes/capillary-postcapillary venule/4 s) to T1 (14.8 [13.2-15.5] leukocytes/capillary-postcapillary venule/4 s) (p < 0.0001). A significant increase in systemic leukocyte count was also detected from 7.4 ± 0.9 × 109/L (preoperative) to 12.4 ± 4.4 × 109/L (postoperative) (p < 0.01). Conclusion: The ability to directly visualize leukocyte-endothelium interaction using IDF imaging facilitates the diagnosis of a systemic inflammatory response after CPB via the identification of rolling leukocytes. Integration of the frame averaging algorithm into the software of handheld vital microscopes may enable the use of microcirculatory leukocyte count as a real-time parameter at the bedside