Intestinal Permeability in Human Nonalcoholic Fatty Liver Disease

A Systematic Review and Meta-analysis

Toon J. I. De Munck; Pan Xu; Harm J. A. Verwijs; Ad A. M. Masclee; Daisy Jonkers; Jef Verbeek; Ger H. Koek

Disclosures

Liver International. 2020;40(12):2906-2916. 

In This Article

Results

Study Selection

Twenty-eight studies were eligible for full text screening. Thirteen of 28 studies matched the criteria and were included in this review. One additional study was identified through reference checking. Excluded studies did not specify alcohol consumption (n = 3), did not use an in vivo IP test (n = 2), did not investigate the association between IP and NAFLD presence or severity (n = 5) or did not include a control group (HC or NAFL) (n = 5). Further details on the selection process can be found in the flowchart (Figure 1).

Figure 1.

Flowchart of the selection process. NAFLD, nonalcoholic fatty liver disease; HC, healthy control; NAFL, nonalcoholic fatty liver (simple steatosis); NASH, nonalcoholic steatohepatitis

Study Characteristics

Nine studies investigated IP with urinary recovery of orally administered molecules (ie sugars or 51Cr-EDTA) (Table 1) and five studies investigated IP with serum zonulin levels (Table 2). Only two studies investigated whole gut permeability by means of 24 h urinary recovery of 51Cr-EDTA or sucralose while all other studies focused on small intestinal permeability.[12,21] In five of fourteen studies BMI was not significantly different between the control group and NAFLD group. However, in only two studies BMI matching of the control group with the NAFLD group (implemented in the study design) was performed.[14,22] Nine (5 adult and 4 pediatric) of fourteen studies used the golden standard, liver biopsy, to diagnose NAFLD, while the five other studies (one adult and four paediatric) used ultrasound. Study characteristics of all included studies are summarized in Table 1 (urinary recovery of orally administered molecules) and Table 2 (serum zonulin).

Quality of Included Studies and Risk of Bias

Table S1 summarizes the quality of all included studies using the adapted NOS. Eight studies had poor quality. Most of these studies scored poorly on the comparability domain, with BMI matching of the control group in only two of the included studies.[14,22] Several factors within all included studies cause heterogeneity across studies. Five different IP tests are used within the 14 studies (zonulin, L/M (5-6 h), L/R (5 h), sucralose (24 h) and 51Cr-EDTA (24 h)). In total, eight studies (5 pediatric and 3 adult) investigated small IP by means of L/M (5-6 h) (7 studies) or L/R (5 h) (1 study), of which data could be extracted and pooled in forest plots. Similarly, data of five studies (three paediatric and two adults) investigating small IP by means of serum zonulin could be pooled. Data of two studies investigated whole gut permeability by means of 24 h urinary collection of sucralose and 51Cr-EDTA were not be pooled because of heterogeneity. Publication bias was not assessed as there were inadequate numbers of included trials in each analysis (less than 10) to properly assess funnel plot asymmetry.[23]

Small Intestinal Permeability in NAFLD Versus HC

Figure 1A shows the quantitative synthesis of the mean L/M or L/R levels of NAFLD subjects vs HC. Seven studies (three adult and four paediatric) comprising a total of 119 NAFLD patients (54 adult and 65 paediatric) and 86 HC (38 adult and 48 paediatric) were included. Overall, NAFLD patients showed an increased small intestinal permeability by means of L/M or L/R (standardized mean difference 0.79 95% CI 0.49–1.08 compared to HC (Figure 2A). The statistical heterogeneity between studies was low (I 2 = 0%). Small intestinal permeability by means of L/M or L/R was increased in both adult and paediatric NAFLD patients compared to HC. However, in the subgroup analysis (adult vs paediatric), the paediatric population showed a significantly higher difference in L/M between the study groups (standardized mean difference 1.09, 95% CI 0.68–1.50 compared to HC).

Figure 2.

Forest plots of small intestinal permeability in NAFLD patients vs healthy controls. (A) Studies using L/M (lactulose to mannitol ratio) or L/R (lactulose to rhamnose ratio) to measure IP; (B) studies using serum zonulin (ng/mL) to measure IP. Including subgroup analysis by age (adult vs paediatric)

Four studies (1 adult and 3 pediatric) comprising a total of 191 NAFLD patients (135 paediatric and 56 adult) and 162 HC (142 pediatric and 20 adult) were included in the quantitative synthesis of the mean serum zonulin levels in NAFLD patients and HC (Figure 1B). Overall, NAFLD patients had an increased small intestinal permeability by means of serum zonulin (standardized mean difference 1.04 ng/mL, 95% CI 0.40–1.68 as compared to HC (Figure 2B). The statistical heterogeneity between studies was high (I 2 = 86%). In the subgroup analysis (Figure 2B), serum zonulin levels were observed to be increased in both adult and paediatric NAFLD patients compared to HC.

Small Intestinal Permeability in NASH vs NAFL

Three studies (one adult and two paediatric) comprising 77 NASH patients (67 paediatric and 10 adult) and 58 NAFL patients (52 paediatric and 6 adult) were included in the quantitative synthesis of the mean L/M levels of NASH vs NAFL patients (Figure 2A). Overall NASH patients had an increased small intestinal permeability by means of L/M (standardized mean difference 0.74, 95% CI 0.17–1.13) compared to NAFL patients (Figure 3A). The statistical heterogeneity between studies was substantial (I 2 = 53%). In the subgroup analysis (Figure 3A), mean L/M was significantly increased in paediatric NASH patients compared to paediatric NAFL patients while this was not the case for adult patients (Figure 3B).

Figure 3.

Forest plots of small intestinal permeability in NASH vs NAFL patients. (A) Studies using L/M (lactulose to mannitol ratio) to measure IP, (B) studies using serum zonulin (ng/mL) to measure IP. Including subgroup analysis by age (adult vs paediatric)

Three studies (two adult and one paediatric) comprising 82 NASH patients (23 paediatric and 59 adult) and 111 NAFL patients (44 paediatric and 67 adult) were included in the quantitative synthesis of the mean serum zonulin levels of NASH vs NAFL patients (Figure 2B). Overall NASH patients had no significantly different serum zonulin levels compared to NAFL patients (standardized mean difference 1.44 ng/mL, 95% CI −0.13–3.00, I 2 = 95%) (Figure 3B). When pooled separately (Figure 3B), in both adult and paediatric patients, no difference in serum zonulin levels between NASH and NAFLD patients was observed.

Whole Gut Permeability in NAFLD

Two adult studies investigated whole gut permeability in adult NAFLD patients by means of 24 h urinary collection of 51CR-EDTA or sucralose.[12,21] Data were not pooled because different markers were used. In the study of Farhadi et al, 24 h sucralose excretion was not significantly different between HC (n = 12), NAFL (n = 6) and NASH (n = 10) patients (Table 1).[21] In the study of Miele et al 24 h 51CR-EDTA excretion was significantly increased in NAFLD patients (n = 35) compared to HC (n = 24) and an increased 51CR-EDTA excretion (median split) was not associated with the presence of NASH.[12]

Association Between Small Intestinal Permeability and NAFLD Severity

Five of the included studies investigated the association between small intestinal permeability and one or more parameters of NAFLD severity (degree of steatosis, fibrosis, ballooning or inflammation) (Table 3). The association between small intestinal permeability and the degree of hepatic steatosis was investigated in three studies (two paediatric and one adult) (Table 3).[14–16] In all studies more advanced hepatic steatosis was associated with an increased small intestinal permeability. To quantify hepatic steatosis all studies used the histological NAFLD activity score (NAS) (Table 1 and Table 2). The association between small intestinal permeability and hepatic fibrosis was investigated in four studies (three pediatric and one adult),[13–16] while three paediatric studies investigated the association with hepatic inflammation and hepatic ballooning (Table 3).[14–16] Only in the study of Giorgio et al, a positive correlation between L/M and the degree of portal inflammation, ballooning and fibrosis was observed in 12 paediatric NASH patients with increased L/M.[15] In all other studies no association between small intestinal permeability and hepatic fibrosis, inflammation or ballooning was observed.

Association Between Whole Gut Permeability and NAFLD Severity

The association between whole gut permeability (24 h 51Cr-EDTA) and NAFLD severity was investigated in one adult study. Miele et al observed that 24 h 51Cr-EDTA excretion was significantly increased in NAFLD patients with moderate to severe steatosis (S2–3) compared to NAFLD patients with minimal or mild steatosis (S1).[12] Furthermore, no difference in degree of hepatic fibrosis, hepatic inflammation or ballooning was observed between patient with normal and increased 24 h 51Cr-EDTA excretion.[12]

Factors That Significantly Correlated With IP in NAFLD Patients

Four of the included studies (one adult and three paediatric) reported significant correlations between small intestinal permeability and clinical factors including anthropometric data and blood biochemical variables (Table 4). Two studies observed a positive correlation between small intestinal permeability and the degree of insulin resistance.[14,22] In addition two other studies observed a positive correlation between small intestinal permeability and systemic LPS levels.[16,24] Other correlations ie with BMI, systolic blood pressure and blood ALT, IL-6, triglycerides, γ-GT and HDL-C levels are only observed in single studies (Table 4).

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