Johanne G. Bloemen

Short chain fatty acids exchange across the gut and liver in humans measured at surgery.

BACKGROUND & AIMS Short chain fatty acids (SCFAs; acetate, propionate and butyrate) are important energy sources for colonocytes and are assumed to play a key role in gut health. Local effects of SCFAs have been investigated, but less is known about whole body metabolism of these SCFAs. The aim of the present study was to quantify the role of the gut and liver in interorgan exchange of SCFAs in humans in vivo. METHODS Twenty-two patients undergoing major upper abdominal surgery were studied. Blood was sampled from a radial artery, the portal and a hepatic vein. Portal, splanchnic and arterial blood flow was measured using intra-operative Duplex ultrasonography. SCFAs were measured on a liquid chromatography system combined with mass spectrometry. RESULTS SCFAs were released by the gut, 34.9 (9.1) micromol kg bodyweight(-1)h(-1). SCFAs uptake by the liver was significant for propionate and butyrate; -5.6 (1.3) and -3.8 (1.6) micromol kg bodyweight(-1)h(-1) (p=0.0002 and p=0.03) respectively and counterbalanced gut release. Liver uptake of acetate was not significant, -5.2 (6.6) micromol kg bodyweight(-1)h(-1) (p=0.434). Splanchnic (i.e., gut+liver) SCFAs release was significant for acetate and propionate, 17.3 (7.3) and 1.2 (0.4) micromol kg bodyweight(-1)h(-1) (p=0.027 and p=0.0038), respectively. Splanchnic release of butyrate was not significantly different from zero (1.9 (1.2) micromol kg bodyweight(-1)h(-1), p=0.129). BMI and previous colonic resection did not affect gut release of SCFAs. CONCLUSION This is the first in vivo study on the role of the gut and liver in SCFAs exchange in humans in vivo. It is shown that intestinal SCFAs release by the gut is equalled by hepatic uptake.

Endogenous formation of Nε-(carboxymethyl)lysine is increased in fatty livers and induces inflammatory markers in an in vitro model of hepatic steatosis

BACKGROUND & AIMS Increased lipid peroxidation and inflammation are major factors in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). A lipoxidation product that could play a role in the induction of hepatic inflammation is N(ε)-(carboxymethyl)lysine (CML). The aim of the present study was to investigate the relationship between steatosis and CML and to study the role of CML in hepatic inflammation. METHODS We included 74 obese individuals, which were categorized into 3 groups according to the grade of hepatic steatosis. CML accumulation in liver biopsies was assessed by immunohistochemistry and plasma CML levels were measured by mass spectrometry. Plasma CML levels were also determined in the hepatic artery, portal, and hepatic vein of 22 individuals, and CML fluxes across the liver were calculated. Hepatocyte cell lines were used to study CML formation during intracellular lipid accumulation and the effect of CML on pro-inflammatory cytokine expression. Gene expression levels of the inflammatory markers were determined in liver biopsies of the obese individuals. RESULTS CML accumulation was significantly associated with the grade of hepatic steatosis, the grade of hepatic inflammation, and gene expression levels of inflammatory markers PAI-1, IL-8, and CRP. Analysis of CML fluxes showed no release/uptake of CML by the liver. Lipid accumulation in hepatocytes, induced by incubation with fatty acids, was associated with increased CML formation and expression of the receptor for advanced glycation endproducts (RAGE), PAI-1, IL-8, IL-6, and CRP. Pyridoxamine and aminoguanidine inhibited the endogenous CML formation and the increased RAGE, PAI-1, IL-8, IL-6, and CRP expression. Incubation of hepatocytes with CML-albumin increased the expression of RAGE, PAI-1, and IL-6, which was inhibited by an antibody against RAGE. CONCLUSIONS Accumulation of CML and a CML-upregulated RAGE-dependent inflammatory response in steatotic livers may play an important role in hepatic steatosis and in the pathogenesis of NAFLD.

Application of liquid chromatography-mass spectrometry to measure short chain fatty acids in blood

A new liquid chromatography-mass spectrometry method is described to determine concentrations of the short chain fatty acids acetic acid, propionic acid and butyric acid (SCFAs) in human blood plasma. The method is based on reversed phase chromatography followed by post-column neutralization of the mobile phase with ammonia and a consecutive measurement of the SCFAs ammonia adducts using negative electro spray ionization. Sample preparation involved simple organic acid deproteinization, resulting in 100% recovery. SCFAs eluted baseline separated within a 25 min run cycle. A linear response was obtained in the range between 0 and 250 micromol/l (R(2) ranged from 0.997 to 0.9999). The limit of detection ranged from 0.05 micromol/l for propionic and butyric acid and 0.1 micromol/l for acetic acid. The method was tested by analyzing plasma of arterial blood, from portal vein and hepatic vein blood from patients undergoing a pylorus-preserving pancreaticoduodenectomy. As expected, the highest SCFA concentrations were found in portal plasma, hepatic vein levels were in between, while arterial concentrations were lowest. This newly developed method is suitable to determine SCFA concentrations in human plasma samples.

Incisional hernias in temporary stoma wounds: a cohort study.

HYPOTHESIS The prevalence of and risk factors for incisional hernias among temporary stoma wounds have implications for clinical practice. DESIGN Retrospective cohort study. SETTING University tertiary care hospital. PATIENTS All adult patients with a stoma closed between January 1, 2000, and August 1, 2004. Of 150 living patients, 111 (74.0%) were included for analysis after follow-up at the outpatient clinic. MAIN OUTCOME MEASURES The main outcome was incisional hernia in a temporary stoma wound, defined as a defect within the musculature and fascia detected by ultrasonographic examination. Risk factors for incisional hernias and the diagnostic validity of clinical symptoms and palpation during the Valsalva maneuver were determined. RESULTS After a median follow-up of 35 months (range, 5-77 months), hernia prevalence was 32.4%. Among patients with a body mass index (calculated as weight in kilograms divided by height in meters squared) of less than 30, hernia prevalence was 25.8%; among patients with a body mass index of 30 or higher, hernia prevalence was 59.1%. Palpation demonstrated the highest sensitivity (58.3%). One in 6 patients had discomfort at the temporary stoma site and no palpable defect but showed an incisional hernia on ultrasonographic examination. Obesity was the sole significant risk factor identified in this study (odds ratio, 5.53; 95% confidence interval, 1.72-17.80). The presence of a stoma in situ for less than 6 months showed a trend toward being a risk factor (odds ratio, 2.38; 95% confidence interval, 0.96-5.99). CONCLUSION Incisional hernias occur in 1 of 3 temporary stoma wounds, and a body mass index of 30 or higher is a risk factor.

Randomized controlled trial analyzing the effect of 15 or 30 min intermittent Pringle maneuver on hepatocellular damage during liver surgery

BACKGROUND & AIMS Aminotransferases are commonly used to determine the optimal duration of ischemic intervals during intermittent Pringle maneuver (IPM). However, they might not be responsive enough to detect small differences in hepatocellular damage. Liver fatty acid-binding protein (L-FABP) has been suggested as a more sensitive marker. This randomized trial aimed to compare hepatocellular injury reflected by L-FABP in patients undergoing liver resection with IPM using 15 or 30 min ischemic intervals. METHODS Twenty patients undergoing liver surgery were randomly assigned to IPM with 15 (15IPM) or 30 (30IPM) minutes ischemic intervals. Ten patients not requiring IPM (noIPM) served as controls. Primary endpoint was hepatocellular injury during liver surgery reflected by systemic L-FABP plasma levels. Between group comparisons were performed using area under the curve and repeated measures two-way ANOVA. RESULTS The IPM groups had similar characteristics. Aminotransferases did not differ significantly between 15IPM and 30IPM at any time point. L-FABP levels rose up to 1853±708 ng/ml in the 15IPM and 3662±1355 ng/ml in the 30IPM group after finishing liver transection and decreased rapidly thereafter. There were no significant differences between 15IPM and 30IPM in cumulative L-FABP level (p=0.378) or L-FABP level at any time point (p=0.149). Blood loss, remnant liver function and morbidity were comparable. CONCLUSIONS IPM with 15 or 30 min ischemic intervals induced similar hepatocellular injury measured by the sensitive marker L-FABP. The present study confirms the results of earlier trials, suggesting that IPM with 30 min ischemic intervals may be used.

Butyrate enemas improve intestinal anastomotic strength in a rat model.

INTRODUCTION: Anastomotic leakage is a common clinical complication with incidences up to 10% to 17% in colorectal surgery, leading to high morbidity and mortality. Butyrate is the product of colonic fermentation of indigestible carbohydrates and is considered beneficial to gastrointestinal healing. The aim of this study was to investigate the effect of intraluminal supplementation of butyrate on colonic anastomotic strength in a rat model. METHODS: Wistar rats were randomly assigned to one of 3 groups (18 animals each). All rats underwent a 1-cm left colonic resection and end-to-end anastomosis with 4 interrupted sutures. Group I underwent no other treatment and served as the control, group II received daily 5 mL of 60 mM sodium butyrate enemas postoperatively, and group III received placebo enemas. On the third or seventh postoperative day, rats (n = 9 per time point) were anesthetized and anastomotic bursting strength was assessed. RESULTS: As a consequence of anastomotic leakage, 3 rats (16.6%) in group I, 1 rat (5.6%) in group II, and 2 rats (11.2%) in group III died. Mean anastomotic bursting pressures at day 3 were not significantly different between groups (53, 64, and 68 mm Hg for group I, II, and III, respectively, P = .777). At day 7, bursting pressures were 118, 225, and 129 mm Hg for groups I, II, and III, respectively (P = .0006). Group II showed an increased mature-to-immature collagen ratio (P = .035). CONCLUSION: Postoperative intestinal butyrate supplementation enhances anastomotic bursting strength in a left-sided partial colonic resection rat model, which can be explained by increased collagen synthesis and maturation.

Short chain fatty acids exchange: Is the cirrhotic, dysfunctional liver still able to clear them?

BACKGROUND & AIMS Prebiotics are increasingly used to improve gut integrity. A presumed mechanism of their beneficial action is the synthesis of short chain fatty acids (SCFA: acetate, propionate and butyrate). High systemic concentrations of propionate and butyrate are toxic and can adversely affect the patient. In physiological situations the liver uses propionate and butyrate for energy metabolism. The aim of the present study was to investigate to which extent patients with liver cirrhosis are still able to metabolize portal derived SCFA in the liver. METHODS Twelve patients with liver cirrhosis and an intrahepatic portosystemic shunt (TIPSS) were studied. Blood was sampled from the femoral artery, portal and hepatic vein. Organ plasma flow was measured. Net release or uptake was calculated by multiplying the arteriovenous differences by plasma flow. SCFA plasma concentrations were measured using LC-MS. RESULTS Arterial concentrations were 124+/-12, 8+/-1 and 10+/-1micromol/l for acetate, propionate and butyrate, respectively. The gut produced 32.5+/-13.0, 4.8+/-1.3 and 6.2+/-2.1micromolkgbw(-1)h(-1) of acetate, propionate and butyrate, respectively. Assuming 70% portosystemic shunting, hepatic uptake of propionate and butyrate was 3.1+/-0.9 and 5.2+/-1.4micromolkgbw(-1)h(-1). Hepatic uptake of acetate was non significant (12.1+/-12.3micromolkgbw(-1)min(-1)). As a consequence of shunting, part of total acetate escaped from the splanchnic bed, which equalled 34.9+/-14.7micromolkgbw(-1)h(-1). CONCLUSION The liver of patients with stable cirrhosis is able to use butyrate and propionate, most likely preventing increased systemic concentrations. This suggests that prebiotics can be administered safely, but monitoring butyrate levels may be advisable in patients with diminished liver function.

Hepatic Uptake of Rectally Administered Butyrate Prevents an Increase in Systemic Butyrate Concentrations in Humans

BACKGROUND Short-chain fatty acids (SCFAs), fermentation products of undigested fibers, are considered beneficial for colonic health. High plasma concentrations are potentially harmful; therefore, information about systemic SCFA clearance is needed before therapeutic use of prebiotics or colonic SCFA administration. OBJECTIVE The aim of this study was to investigate the effect of rectal butyrate administration on SCFA interorgan exchange. METHODS Twelve patients (7 men; age: 66.4 ± 2.0 y; BMI 24.5 ± 1.4 kg/m(2)) undergoing upper abdominal surgery participated in this randomized placebo-controlled trial. During surgery, 1 group received a butyrate enema (100 mmol sodium butyrate/L; 60 mL; n = 7), and the other group a placebo (140 mmol 0.9% NaCl/L; 60 mL; n = 5). Before and 5, 15, and 30 min after administration, blood samples were taken from the radial artery, hepatic vein, and portal vein. Plasma SCFA concentrations were analyzed, and fluxes from portal-drained viscera, liver, and splanchnic area were calculated and used for the calculation of the incremental area under the curve (iAUC) over a 30-min period. RESULTS Rectal butyrate administration led to higher portal butyrate concentrations at 5 min compared with placebo (92.2 ± 27.0 μmol/L vs. 14.3 ± 3.4 μmol/L, respectively; P < 0.01). In the butyrate-treated group, iAUCs of gut release (282.8 ± 133.8 μmol/kg BW · 0.5 h) and liver uptake (-293.7 ± 136.0 μmol/kg BW · 0.5 h) of butyrate were greater than in the placebo group [-16.6 ± 13.4 μmol/kg BW · 0.5 h (gut release) and 16.0 ± 13.8 μmol/kg BW · 0.5 h (liver uptake); P = 0.01 and P < 0.05, respectively]. As a result, splanchnic butyrate release did not differ between groups. CONCLUSION After colonic butyrate administration, splanchnic butyrate release was prevented in patients undergoing upper abdominal surgery. These observations imply that therapeutic colonic SCFA administration at this dose is safe. The trial was registered at clinicaltrials.gov as NCT02271802.

Liver manipulation during liver surgery in humans is associated with hepatocellular damage and hepatic inflammation.

Manipulation of the liver during liver surgery results in profound hepatocellular damage. Experimental data show that mobilization‐induced hepatocellular damage is related to hepatic inflammation. To date, information on this link in humans is lacking. As it is possible to modulate inflammation, it is clinically relevant to unravel this relationship.

Gut and liver handling of interleukin-6 during liver resection in man

BACKGROUND Plasma interleukin-6 (IL-6) levels increase during liver resection. The source of this IL-6 is hitherto unclear. It has been demonstrated that the hepatosplanchnic area takes up IL-6 but the role of the gut and liver is unknown. The aim of the present study was to investigate the role of the gut and liver in IL-6 homeostasis during liver surgery. METHODS Before and after partial hepatectomy, IL-6 was measured in blood sampled from the radial artery, and the hepatic and portal vein. Blood flow was measured to assess IL-6 fluxes (flow times AV-differences) across the gut, liver and hepatosplanchnic area. RESULTS In 22 patients undergoing liver resection, IL-6 release from the gut after transection was 90.9 (30.1) ng/min (P < 0.001), whereas net IL-6 uptake by the liver equalled 83.4 (41.7) ng/min (P < 0.01). Overall hepatosplanchnic flux was 7.3 (43.5) ng/min after transection and did not differ significantly from zero. Overall hepatosplanchnic flux was 87.8 (41.5) ng/min in the major resection group and -59.8 (67.5) ng/min in the minor resection group (P < 0.05). DISCUSSION The gut releases IL-6 and the liver takes up IL-6 before and after liver resection. The loss of IL-6 uptake as a result of a small functional remnant liver could lead to higher IL-6 levels after surgery.