Mucosal inflammation and injury in the intestinal segment orad to a strangulating lesion has been reported by several authors.1–5 Pathological changes in this section of intestine may contribute to postoperative complications, such as motility disorders or adhesions, and thereby affect the survival of the horse.6,7 These changes may not be visible macroscopically, which can pose a problem during colic surgery when determining which intestinal segment necessitates resection.1
Pathohistological examination is still considered the gold standard for the diagnosis of intestinal injury.4,8 However, this is not available as instant diagnostic procedure during surgery. Another issue for the implementation of histology for intraoperative decision making is that the correlation between the extent of histologically detectable injury in the orad segment and the occurrence of postoperative complications has not been established. The assessment of intestinal microperfusion has been investigated as a possible diagnostic aid to estimate the degree of tissue damage. However, most techniques used to quantify the intestinal blood flow, such as surface oximetry, fluorescence and Doppler ultrasound, have focused on ischemic or reperfused intestine and were not applied to the orad segment.9–11 Laser Doppler flowmetry (LDF) is a technique based on detecting the Doppler shift of laser light on moving red blood cells, which can be combined with white light spectrophotometry to determine the oxygen saturation and hemoglobin content of the tissue.12 This has been applied in strangulated and orad segments in equine colic cases, reporting reduced microperfusion in the obstructed segment but not in the orad segment.13 Furthermore, the ability of LDF to detect ischemic areas in strangulated intestine and to select the proper level of resection was demonstrated in human medicine.14,15 To the authors’ knowledge, no previous studies have documented the blood flow of the orad segment of naturally occurring strangulating diseases combined with histology and postoperative outcome in horses or any other animal species.
Therefore, the aim of this study was to determine the diagnostic value and practicability of LDF spectrophotometry (LDFS) for the assessment of injury in the segment orad to a small intestinal strangulation in naturally occurring disease. The first objective was to describe the microperfusion as well as the mucosal injury, inflammation, and hypoxia inducible factor 1 alpha (HIF-1α) immunoreactivity of the orad intestine during colic surgery. We hypothesized that the orad segment would have lower microperfusion values than the previously published reference values and that the orad segment would show histological injury. The second objective was to determine if cases with reduced microperfusion were more likely to suffer from postoperative reflux (POR) or nonsurvival. We hypothesized that horses with low tissue oxygen saturation (tSO2) would have a higher incidence of POR and nonsurvival. The third objective was to compare the microperfusion between the segments with and without pathohistological injury. We hypothesized that the segments with mucosal injury would have lower tSO2 and tissue blood flow (tBF).
Methods
Animals
Ethical approval was obtained from the animal welfare officer of the University of Veterinary Medicine Hannover Foundation, Germany, and the ethics committee of the responsible German federal state authority in accordance with the German Animal Welfare Law (Lower Saxony State Office for Consumer Protection and Food Safety, LAVES 33.8-42502-05-19A452). For this prospective clinical trial, client-owned horses undergoing a laparotomy for acute colic were enrolled in the study. The decision to perform colic surgery was based on clinical ultrasonographic and laboratory findings and was independent of the study. Horses were eligible for enrollment in the study if a strangulating small intestinal lesion was diagnosed during colic surgery. Cases were included in the study if there was an opportunity to conduct the LDFS measurements in the segment orad to the strangulated segment and/or to take a histology sample from this section. Consent was obtained from owners of all horses included in the study. Cases were excluded if the measurement would cause a significant prolongation of surgery time or if any delay was considered unacceptable due to the situation in surgery or the anesthetic stability of the horse. Other reasons for exclusion were personnel constraints and technical issues.
Laparotomy and intraoperative measurements
A routine ventral midline laparotomy was performed in dorsal recumbency under general anesthesia. Anesthetic induction and maintenance as well as monitoring were performed in accordance with the routine protocol, including direct blood arterial blood pressure measurement in the facial artery and arterial blood gas analysis. Following release of the strangulation and exteriorization of the affected intestinal segment, the LDFS measurement was performed prior to manual decompression of the intestine. This was done utilizing an LDFS probe with a penetration depth of 2.5 mm (O2C with LF-2 probe; Lea Medizintechnik GmbH) covered with a transparent sterile ultrasound sleeve (Flexasoft; Udo Heisig GmbH) that was placed on the antimesenteric side of the intestine 0.5 to 1 m orad to the strangulation for a minimum of 30 seconds. The relative tBF in arbitrary units (AU) was measured by detection of the Doppler shift of a < 30-mW, 830-nm laser, and tSO2 (%) and the relative hemoglobin (tHB in AU) was measured by white light spectrophotometry (500 to 850 nm). The LDFS measurements were recorded with 50 Hz, and the output summary with averaged values for each 2 seconds was used for further analysis. To exclude motion artifacts, only the lower 50% of the values of each measurement were included in the final analysis.
The decision regarding the need for intestinal resection and its extent was determined by the attending surgeon based on clinical assessment of tissue vitality using a previously published score.16 If intestinal resection was deemed necessary, intestinal biopsies were taken from the orad border of the resected segment. In cases necessitating resection of large segments of small intestine (> 60% to 75%) and/or a jejuno-cecostomy, some owners elected to discontinue treatment. In these cases, the intestinal biopsies were taken immediately postmortem.
Following surgery, the relevant case characteristics were collected from the hospital records. The intraoperative findings (diagnosis, localization, and length of affected segment; the clinical viability score, wall thickness, location, and timing of measurement; treatment) were documented in the standardized surgery report and by use of an additional short questionnaire. The occurrence of POR, defined as > 1 L/h during > 24 hours, was recorded as well as other complications. Furthermore, survival to hospital discharge and findings at relaparotomy or postmortem examination were documented during the progression of the case.
Histology
The intestinal tissue was carefully rinsed in isotonic saline solution, and the biopsy segment was fixed in 4% formaldehyde solution for 24 to 36 hours and then embedded in paraffin. Following routine processing, the sections were stained with H&E. Two sections per sample were assessed by one observer using light microscopy (AXIO Scope.A1; Carl Zeiss Vision GmbH) at a 400-fold magnification. The mucosal damage was quantified using a modified semiquantitative mucosal injury score (MIS)17–19: grade 0, normal mucosal villi; grade 1, slight separation of epithelial cells from the lamina propria mucosae at the tip of the villus (Gruenhagen’s space); grade 2, extension of subepithelial space plus or minus loss of epithelial cells from the tip of the villus; grade 3, extension of the subepithelial space with epithelial lifting down the sides of the villi exposing one-fourth to three-fourths of the lamina propria; grade 4, complete or nearly complete separation of epithelium from lamina propria to the villus base (denuded villi), crypts and basal membrane intact; grade 5, partial loss of villus architecture (of the lamina propria) and partial necrosis of the crypts; and grade 6, nearly complete to complete loss of lamina propria architecture with severe necrosis of the crypts. Furthermore, a hemorrhage score (HS) was used to grade the extent of hemorrhage in all intestinal layers individually20: grade 0, none; grade 1, few extravascular individual erythrocytes; grade 2, increased number of individual extravascular erythrocytes; grade 3, moderate hemorrhage and erythrocyte clumping at a few sites; grade 4, multifocal severe clumping of erythrocytes, tissue structure may be affected but is still recognizable; and grade 5, massive hemorrhage obscuring the majority of normal tissue architecture.
Immunohistochemical staining for cytosolic calprotectin (monoclonal mouse anti-human myeloid/histiocyte antigen, clone MAC 387, catalog #M0747; immunoglobulin G concentration, 375 mg/L; Dako Deutschland GmbH) was performed to quantify the inflammatory cells as previously described.21 The cells with clear immunoreactivity were counted in 5 adjoined high-power fields (HPF; 400-fold magnification) in each intestinal layer and expressed as positive cells per 5 HPF. If > 50 positive cells were present in each HPF, a count of 250 cells per 5 HPF was noted for that intestinal layer.
Furthermore, the slides were immunohistochemically stained for HIF-1α (polyclonal rabbit anti-human HIF-1α antibody; catalog #NB100–134; immunoglobulin G concentration, 1.0 mg/mL; Novus Biologicals LLC).22 The immunoreactivity of enterocytes was graded for the staining intensity of the cytoplasm and the nucleus in the crypts and villi using the following score: grade 0, no staining; grade 1, weak staining (staining hardly visible); grade 2, mild staining (light brown); grade 3, moderate staining (medium brown); and grade 4, intense staining (dark brown). This was performed by one observer at fixed microscopic settings, and microscopic photographs were used as color references. The total immunoreactivity score was the sum of the cytoplasm and nucleus score of both the villi and the crypts.
Data analysis
An a priori power analysis (G-Power, version 3.1.9.6; Heinrich Heine Universität) yielded a sample size of 7 horses per group to detect a difference in tSO2 or tBF of 50 with an SD of 30 between the horses with or without mucosal injury, with a power of 0.8 and an alpha of .05. For graph design and statistical analysis, commercially available software was used (Graphpad Prism, version 9.4.1; Graphpad Software Inc). To test for (log-)normal distribution, a Shapiro-Wilks test was performed. Variance homogeneity was assessed with Levene’s test and the visual assessment of the homoscedasticity plots. Normally distributed variables were expressed as mean ± SD, and the other variables as median (minimum – maximum). The histology scores were displayed as frequency distribution. The tSO2 and tHB were not (log)normally distributed, whereas tBF showed a normal distribution.
To compare the LDFS between segments with and without mucosal injury (MIS ≥ 1 and MIS = 0, respectively), tSO2 and tHB were compared using a Mann-Whitney U test, and the tBF was compared using a two-samples t test. The number of horses with POR and the number of horses that survived were compared between groups using Fisher’s exact test. To test for a correlation between different variables (ie, arterial and tissue SO2 as well as LDFS measurements and histology findings), the nonparametric Spearman correlation coefficient was calculated. P values of < .05 were considered significant.
Results
Intraoperative findings and outcome
Thirty-one horses were included during the study period (Figure 1). Fifteen horses had a strangulating lipoma, and 7 horses had an entrapment in the epiploic foramen. Furthermore, there were 4 horses with a mesenteric hernia, 3 with a volvulus nodosus, 1 with an omental hernia, and 1 with herniation through the gastrosplenic ligament. In cases necessitating intestinal resection and anastomosis, the resection margins were judged by the surgeon to be viable. Apart from mild serosal petechiae or mild wall thickening, no gross abnormalities of the intestinal wall of the orad segment were observed. Length of resection ranged from 0.5 to 9 meters.
Flow diagram depicting the cases that were included in this prospective study, including treatment and outcome. LDFS = laser Doppler flowmetry and spectrophotometry.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Flow diagram depicting the cases that were included in this prospective study, including treatment and outcome. LDFS = laser Doppler flowmetry and spectrophotometry.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Flow diagram depicting the cases that were included in this prospective study, including treatment and outcome. LDFS = laser Doppler flowmetry and spectrophotometry.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Postoperatively, 2 of 3 horses that did not undergo intestinal resection developed POR. All 4 horses with a jejuno-ileostomy had POR, of which 3 did not survive due to obstruction at the anastomoses, adhesions, or stricture of the orad jejunum in 1 case. Two of 3 horses with a jejuno-cecostomy developed POR, which resolved with conservative treatment. Three of 12 horses with jejuno-jejunostomy had POR, of which 1 resolved with conservative treatment, and 2 had to be euthanized because of persistent reflux and colic. At necropsy, necrosis and/or adhesions of the remaining intestine could be found as cause for this clinical deterioration.
LDFS measurements
The microperfusion in the orad segment showed a high individual variability, with the majority of horses exhibiting lower tSO2 and tBF as well as a higher tHB compared to the published reference values (Table 1).
Intestinal microperfusion in the intestinal segment orad to naturally occurring small intestinal strangulations.
| tSO2 (%)a | tHB (AU)a | tBF (AU)b | |
|---|---|---|---|
| Total, including horses without histology (n = 18) | 50.2 (4.4–95.0) | 87.7 (43.3–106.0) | 147.3 ± 123.3 |
| Segments without mucosal injury (n = 7) | 79.0 (16.9–90.2) | 89.1 (43.3–106.3) | 155.7 ± 116 |
| Segments with mucosal injury (n = 8) | 30.4 (4.4–87.5) | 88.9 (79.2–102.4) | 134.7 ± 155.3 |
| Reference valuesc | 81.0 ± 13 | 68.4 ± 11 | 246.1 ± 73 |
Looking at the individual values of tSO2, there were 9 horses with tSO2 < 35% and 9 horses with tSO2 > 69% (Figure 2). Of the horses with tSO2 < 35%, 3 of 9 were euthanized intraoperatively, and from the remaining 6, only 2 survived, which both suffered from POR. Of the horses with tSO2 > 69%, 3 of 9 were euthanized intraoperatively, and the remaining 6 horses all survived. The occurrence of POR differed significantly between horses with low and high tSO2 (6/6 and 1/6, respectively; P = .02). The difference in survival between horses with low and high tSO2 did not reach statistical significance (2/6 and 6/6, respectively; P = .06). The 4 horses with low tSO2 that were euthanized during hospitalization included 2 jejuno-jejunostomies, 1 jejuno-ileostomy, and 1 horse with repositioning only. In 1 horse with a jejuno-jejunostomy, the segment orad to the anastomosis was found to be necrotic. The horse with the jejuno-ileostomy exhibited an impaction at the anastomosis treated by a jejuno-caecal bypass during the first relaparotomy, but POR persisted. At the second relaparotomy, multifocal adhesions were found, and the horse was euthanized. The remaining 2 horses had hypomotile small intestines without clear macroscopic intestinal wall changes or obvious signs of anastomotic dysfunction at relaparotomy or necropsy.
Individual value plot of the tissue oxygen saturation of the intestinal segment orad to a strangulating lesion, depicting the occurrence of postoperative reflux and the survival to discharge in 18 surgical colic cases.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Individual value plot of the tissue oxygen saturation of the intestinal segment orad to a strangulating lesion, depicting the occurrence of postoperative reflux and the survival to discharge in 18 surgical colic cases.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Individual value plot of the tissue oxygen saturation of the intestinal segment orad to a strangulating lesion, depicting the occurrence of postoperative reflux and the survival to discharge in 18 surgical colic cases.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Arterial blood gas measurements taken from the facial artery within 10 minutes prior to or after the LDFS measurement yielded a mean PaO2 of 128.9 ± 71 mm Hg (range, 54 to 523 mm Hg) and a mean arterial oxygen saturation (aSO2) of 96.2 ± 0.3% (range, 87.5% to 99.9%). The mean arterial pressure (MAP) was 88.6 ± 18 mm Hg (range, 61 to 104 mm Hg). There was no significant correlation between the aSO2 and tSO2 (r = 0.04; CI, –0.44 to 0.51; P = .9) or between the MAP and tBF (r = –0.14; CI, –0.58 to 0.37; P = .6).
Pathohistology
Most horses had a grade 0 for the MIS and HS (Figure 3), and none exhibited crypt necrosis or destruction of the lamina propria mucosae. Of the 19 horses recovered from anesthesia with available intestinal biopsies, 6 horses had significant epithelial separation (grade 3 to 4) in the orad segment, and all of these horses had POR. The remaining 13 horses had no epithelial separation at all (grade 0), of which 3 had POR. This difference in the occurrence of POR between these groups was significant (P = .003). No pathomorphological changes were seen in the tunica muscularis nor in the submucosal and myenteric plexus.
Histogram depicting the mucosal injury of 28 small intestinal segments orad to naturally occurring strangulations by use of a semiquantitative mucosal injury score (MIS; grades 0 to 6) and a hemorrhage score (grades 0 to 5) of the different intestinal layers (right panel). Grade 0 represents no injury or hemorrhage. These results include the cases that were euthanized during surgery.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Histogram depicting the mucosal injury of 28 small intestinal segments orad to naturally occurring strangulations by use of a semiquantitative mucosal injury score (MIS; grades 0 to 6) and a hemorrhage score (grades 0 to 5) of the different intestinal layers (right panel). Grade 0 represents no injury or hemorrhage. These results include the cases that were euthanized during surgery.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Histogram depicting the mucosal injury of 28 small intestinal segments orad to naturally occurring strangulations by use of a semiquantitative mucosal injury score (MIS; grades 0 to 6) and a hemorrhage score (grades 0 to 5) of the different intestinal layers (right panel). Grade 0 represents no injury or hemorrhage. These results include the cases that were euthanized during surgery.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Looking at the microperfusion in the segments with and without histologically detectable injury, the tSO2 of the noninjured segments was subjectively higher than that of the injured segments (Table 1), yet this was not statistically significant (median difference, 48.6%; P = .4) (Table 1). The tBF also did not differ between the injured and noninjured segments (mean difference, 21.0 AU; P = .8). Furthermore, there was no significant correlation between the tHB and the cumulative HS (r = 0.15; CI, –0.39 to 0.61; P = .6).
There was a great variation in the total count calprotectin-positive cells, with most horses exhibiting only small numbers of positive cells, with a median of 49.5 (6 to 951) cells per 5 HPF. Most positive cells were present in the mucosa, with 13 (3 to 250) cells per 5 HPF, followed by the serosa, muscularis, and submucosa (9.5 [1 to 250], 4 [1 to 250], and 2 [1 to 250], respectively). There was no difference in total calprotectin-positive cell count between the horses with and without POR (115 [17 to 950] and 52 [7 to 544] cells per 5 HPF, respectively). Furthermore, there was no significant correlation between the total cell count and the tSO2 (r = –0.23; CI, –0.70 to 0.28; P = .3) or tBF (r = –0.15; CI, –0.63 to 0.41; P = .6).
The mucosa of all samples stained positive for HIF-1α, with stronger immunoreactivity in the crypts than in the villi (Figure 4). There was no significant correlation between the total HIF immunoreactivity score and the tSO2 (r = 0.13; CI, –0.43 to 0.61; P = .7).
Histogram depicting the immunoreactivity for hypoxia inducible factor 1 alpha (HIF1α) of small intestinal segments orad to naturally occurring strangulations. Grade 0, no staining; grade 1, weak staining; grade 2, mild staining; grade 3, moderate staining; and grade 4, intense staining.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Histogram depicting the immunoreactivity for hypoxia inducible factor 1 alpha (HIF1α) of small intestinal segments orad to naturally occurring strangulations. Grade 0, no staining; grade 1, weak staining; grade 2, mild staining; grade 3, moderate staining; and grade 4, intense staining.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Histogram depicting the immunoreactivity for hypoxia inducible factor 1 alpha (HIF1α) of small intestinal segments orad to naturally occurring strangulations. Grade 0, no staining; grade 1, weak staining; grade 2, mild staining; grade 3, moderate staining; and grade 4, intense staining.
Citation: American Journal of Veterinary Research 2024; 10.2460/ajvr.23.12.0286
Discussion
This study describes the microperfusion of intestine orad to naturally occurring strangulations, showing that the tSO2, tHB, and tBF could be measured by LDFS during colic surgery. The main findings were that the microperfusion in this segment was lower than reported in horses without gastrointestinal disease in a previous report.23 Furthermore, around half of the intestinal segments also showed morphological changes or hemorrhage of the mucosa, thereby partially confirming the first hypothesis that the orad segment would have decreased microperfusion and would show histological injury. Horses with low tSO2 were more likely to suffer from POR than horses with high tSO2. The difference in survival rate to discharge did not reach statistical significance, consequently only partially accepting the second hypothesis. The tSO2 and tBF values of the segments without histological injury were higher than those of the injured segments, yet this was not statistically significant, possibly due to the high variability of the data. Furthermore, there was no correlation between the tHB and the tissue hemorrhage or between tSO2 and HIF-1α immunoreactivity. Therefore, the third hypothesis stating that segments with mucosal injury would have lower microperfusion measurements was rejected.
Even though the orad segment is not strangulated directly, the current study found low tSO2 levels in half of the horses. Distention orad to the strangulating obstruction may explain the decreased blood flow in the orad segment, considering that previous experimental in vivo studies have shown that intraluminal distention negatively affects the mesenteric blood flow24 and decreases the number of perfused blood vessels in the seromuscular layer.25 Other possibilities could include excessive tension on the mesentery due to displacement or distention of the strangulated segment. Furthermore, decreased intestinal perfusion can also result from the use of anesthetics, such as isoflurane, or generalized hypovolemia and hypoxemia.26,27 However, this is not very likely in the horses of the current study considering that the MAP and aSO2 were above critical limits and that there was no correlation between the aSO2 and tSO2 or between the MAP and tBF. Contrary to the results of the current study, another study13 investigating the use of LDFS during colic surgery did not report reduced microperfusion of the orad segment. This discrepancy might be caused by differences in the degree of distention or the amount of bowel manipulation, considering blood flow has been shown to increase significantly during decompression of the intestine.24 The penetration depth of 2.5 mm of the probe should also be considered in the interpretation of the results, and wall thickness may also account for differences between measurements. In the current study, wall thickness of the orad segment was not measured specifically, but no gross wall thickening was recorded. Considering that the normal wall thickness of the jejunum is not expected to exceed 2 to 3 mm,28,29 the majority of the mucosa is most likely included in the measurement.
An important finding was that horses with low tSO2 were more likely to suffer from POR compared to horses with high tSO2. Microperfusion of the orad segment following strangulation has not been investigated in relation to the outcome of naturally occurring strangulating lesions. However, there are some parallels between measuring blood flow in the orad segment and assessing blood flow at an anastomosis. For the latter, it has been shown that a decreased intraoperative colonic tSO2 value of ≤ 90% measured by pulse oximetry was associated with the occurrence of anastomotic leakage in colorectal surgery in humans.30 Furthermore, a positive correlation has been found between a decrease in blood flow in the rectal stump measured by LDF and anastomotic leakage rate in humans.31 An exact comparison of these values with the measurements of the current study should be done with caution as this differs in timing, bowel segment, manipulation prior to measurement, and the incorporation of tissue into a suture or staple line. However, it concerns approximately the same location in relation to the strangulation, and it illustrates the potential value of blood flow assessment in this segment in predicting the postoperative outcome.
Histology of the orad segment revealed significant epithelial separation in 6 of 19 horses that recovered from anesthesia with available intestinal biopsies. This is in line with previous studies1–4,32 reporting mucosal injury in this segment. In the current study, none of these 6 sections had an HS of > 2 in the serosa and muscularis. This may explain why none of the intestinal segments was clinically judged to be injured, again confirming that visual assessment of the serosal aspect is not representative for the detection of mucosal injury. All horses with mucosal injury had POR, which was significantly less likely to occur in horses that had no epithelial separation. Even though mucosal injury may not directly be accountable for motility issues, it shows that the intestinal segment has undergone considerable ischemia, which may also elicit pathological changes in the other intestinal layers that may not be seen at the time of sampling or by morphological assessment of H&E-stained biopsies alone but could result in POR later on. The tSO2 and tBF values of the segments without histological injury were higher than those of the injured segments, yet this was not statistically significant. Even though a power analysis was performed, the high variability between the individual cases and the many factors that can influence the intestinal blood flow (measurement) could have caused smaller differences to go undetected.
Most horses only had small numbers of inflammatory cells in the intestinal wall of the orad segment. Furthermore, there was no difference in total calprotectin-positive cell count between the horses with and without POR. This could be interpreted as contradictory to the theory that inflammation is a main cause of POI. However, it should be considered that the samples were taken almost immediately following resolution of ischemia, whereas it is known that the neutrophil count increases up to 24 hours following manipulation.33,34 Therefore, the neutrophil influx was most likely not at its peak at the time of sampling in the current study. Alternatively, the POR in these cases might not be caused by a generalized functional ileus in the orad intestine, yet it could also result from a localized functional or mechanical ileus at the level of the anastomosis. The mucosa of all samples stained positive for HIF-1α without any intense immunoreactivity, and there was no significant correlation between the total HIF-1α immunoreactivity score and the tSO2. This may be explained by the degradation of HIF-1α due to oxygen exposure during sampling and questions the use of HIF-1α as a measure of tissue hypoxia for the current sampling conditions.
One of the main limitations of the study is that only 1 LDFS measurement was performed peripheral to the strangulating lesion. It would have been useful to assess the intestine further orad or at the aborad border as this could have provided more information on the role of distention or of the distance to the strangulated segment. Furthermore, assessing blood flow before and after reduction of the strangulation as well as before and after decompression or resection of the intestine could have provided more insight into these topics. These additional measurements were not performed due to time constraints as measurements were performed during > 30 seconds to increase the reliability of the data considering the risk of motion artifacts and operator differences. Therefore, obtaining data from different locations at different time points would have increased the surgery time, which was deemed to be unacceptable in this clinical study that already included several measurements of the strangulated segment.35 Another important limitation of the study was the limited number of cases, precluding the calculation of an exact cut of values or negative and positive predictive values and possibly limiting the detection of smaller differences between the groups. Therefore, this should be seen as an exploratory study investigating the utility of this device, and the reported values may be indicative of intestinal hypoperfusion but do not represent set reference ranges. Furthermore, the cases that did not undergo intestinal resection did not have intestinal biopsies taken, excluding these cases from the association between microperfusion and histology. Another point of interest would have been to assess the intraluminal pressure in relation to the microperfusion, which was not included as this would mean puncturing the intestine. Additional surgical manipulation of intestines that were to remain in situ was considered to be unacceptable in these clinical cases. Another limitation is that the microperfusion may have been altered following exteriorization of the intestine compared to the situation intra-abdominally. However, if the intestine was to remain in the abdomen during the measurement, it would not have been possible to reliably ascertain which segment should be measured as orad segment. Also, the duration of colic could not be included because when evaluating the history data, it became apparent that in most cases the horses had been found with colic after they had been unobserved for a longer period. Therefore, the factor of duration of hypoperfusion remains unclear in the interpretation of the results.
In conclusion, the tSO2 of the intestine orad to a strangulation was lower than previously reported in healthy horses. Furthermore, horses with low tSO2 were more likely to suffer from POR and did not survive until discharge. The findings should be interpreted in the light of the numerous limitations inherent to a clinical study. Nevertheless, the results indicate that intraoperative tSO2 measured by white light spectrophotometry of the orad segment may aid in predicting postoperative issues. Furthermore, it confirms the significance of this segment for postoperative outcome and highlights the limitations of its clinical assessment. This was also confirmed by the finding that mucosal injury could be detected in segments that appeared normal macroscopically and that horses with intestinal injury were more likely to suffer from POR. The blood flow measured by LDF did not exhibit such clear differences, suggesting that tSO2 measurements might be more useful for the viability assessment of the intestine peripheral to a strangulation lesion. The data set of the current study is too small to determine exact cutoff values for resection margins, and future studies including more cases with multiple measurements are necessary to establish the reference ranges needed for its reliable use during colic surgery.
Acknowledgments
The authors would like to thank all involved employees of the clinic for horses who contributed to the execution of the study. Furthermore, the authors are grateful to Kerstin Rohn for her expert support in tissue processing and immunohistochemistry.
Disclosures
Sabine Kästner is a member of the AJVR Scientific Review Board, but was not involved in the editorial evaluation of or decision to accept this article for publication.
No AI-assisted technologies were used in the generation of this manuscript.
Funding
The research was funded by the authors’ departments.
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