Unveiling the Ghrelin and Obestatin Roles in Inflammatory Bowel Diseases: A Systematic Review and Meta-Analysis Assessing Their Pathogenic Implications and Biomarker Utility

Abstract

Background

Inflammatory bowel disease (IBD), pathologically known as chronic inflammation of the gastrointestinal tract, is among the diseases with a high burden worldwide. Ghrelin and obestatin, as adipocytokines mainly in adipose tissues, are involved in immune responses and inflammatory pathways. Studies have assessed the circulatory ghrelin levels in patients with IBD. Herein, we aim to pool these studies through systematic review and meta-analysis.

Methods

Four international databases, PubMed, Embase, Scopus, and the Web of Science were systematically searched for studies assessing ghrelin or obestatin levels in patients with IBD (either Crohn’s disease [CD] or ulcerative colitis [UC]) in active phase or in remission. Random-effects meta-analysis was conducted in order to calculate the pooled estimate using the standardized mean difference (SMD) and 95% confidence interval (CI).

Results

Nineteen studies were included in our systematic review, comprising 1064 patients with IBD (476 UC and 588 CD). A meta-analysis of 11 studies for comparison of active and quiescent disease showed that patients with active IBD had significantly higher levels of ghrelin (SMD, 0.70; 95% CI, 0.06 to 1.34; P = .03). However, in separate analyses for UC or CD, no such difference was observed (SMD, 1.30; 95% CI, −0.28 to 2.88, P = .11; and SMD, 0.80; 95% CI, −0.41 to 2.01; P = .20, respectively). No significant difference was also observed in ghrelin levels between patients with active IBD and healthy control subjects. Obestatin levels also were not different between patients with active disease and those in remission (SMD, 0.31; 95% CI, −0.05 to 0.68; P = .09). On the other hand, the obestatin/ghrelin ratio was significantly lower in patients with active IBD (SMD, −1.90; 95% CI, −2.45 to −1.35; P < .01).

Conclusions

Our results demonstrate that IBD patients with active disease have higher levels of ghrelin, which needs to be confirmed in future studies. Also, the obestatin/ghrelin ratio might be a promising biomarker for the assessment of disease activity.

Introduction

Inflammatory bowel diseases (IBDs), comprising ulcerative colitis (UC) and Crohn’s disease (CD), are caused by the chronic inflammation of the gastrointestinal tract. These diseases result from an exaggerated immune response against specific antigens or environmental factors in genetically susceptible individuals. IBDs manifest through alternating periods of remission and activation.¹ The global prevalence of IBDs is estimated at approximately 6.8 million cases, with higher occurrence in high-income nations and an increasing trend in developing countries.²

The diagnosis of IBDs is confirmed through endoscopic findings, histology, clinical features, and imaging.³ Due to the invasive and costly nature of current diagnostic methods for IBDs, significant efforts have been made to identify noninvasive, accurate, and cost-effective biomarkers for diagnosis and disease monitoring for prognosis. Unfortunately, despite ongoing efforts, no biomarker with these desired characteristics has been discovered yet. C-reactive protein and fecal calprotectin are recognized biomarkers associated with IBD activity; however, they lack sensitivity.⁴ Consequently, the search for ideal biomarkers for monitoring the disease activity and flare-ups in IBD continues. Previous studies have explored various markers, including fecal lactoferrin and calprotectin, interleukin (IL)-6, elastase, and adipocytokines.

Adipocytokines are a subset of cytokines primarily, although not exclusively, synthesized by white adipose tissue. White adipose tissue comprises a major component of the body’s adipose tissue and, in addition to energy regulation, exerts crucial roles in immunological and metabolic processes. The essential adipocytokines identified so far are leptin, resistin, adiponectin, obestatin, visfatin, and ghrelin. Moreover, adipose tissue can contribute to inflammatory pathways by producing IL-6, tumor necrosis factor α (TNF- α), and plasminogen activator inhibitor type 1.⁵

Ghrelin, a 28-amino-acid peptide hormone, acts as an endogenous ligand of the growth hormone secretagogue receptor, and its initial discovery in 1990 revealed its involvement in growth hormone secretion, appetite stimulation, gastrointestinal motility, glucose tolerance, and energy homeostasis. Apart from these well-established roles, ghrelin also participates in immune responses and exhibits significant anti-inflammatory activities by reducing proinflammatory cytokines like IL-6, IL-1α, IL-1β, and TNF-α. High ghrelin levels have been observed in variety of autoimmune diseases, such as rheumatoid arthritis, autoimmune thyroiditis, multiple sclerosis, and type 1 diabetes, implicating its association with the underlying pathogenesis of these conditions.⁶ Ghrelin has also been investigated in autoimmune gastrointestinal diseases such as celiac disease and IBDs. The association between ghrelin and IBDs was first examined in 2006 by Karmiris et al,⁷ who demonstrated increased ghrelin levels in IBD patients compared with healthy control subjects. Some subsequent studies have reported similar results,^8,9 while others have not found any differences in ghrelin levels between IBD patients (either active or in remission) and healthy control subjects.¹⁰

Ghomraoui et al’s¹¹ study demonstrated that ghrelin levels are lower in IBD patients than in control subjects, and active IBD cases have lower levels than those in remission. Furthermore, a small number of studies have assessed the effects of anti-TNF therapy on ghrelin levels, with one study indicating a reduction in ghrelin levels following treatment, while another study reported an increase.¹² Moreover, patients with IBD are at an increased risk of osteopenia and osteoporosis due to increased absorption and decreased retention of bone minerals compared with the normal population. A study by Koutroubakis et al¹³ demonstrated an association between increased ghrelin levels in IBD patients and reduced bone mineral density. As a result, there is currently no consensus regarding the relationship between ghrelin and IBD.

Another marker investigated in IBD, although to a lesser extent than ghrelin, is obestatin. Obestatin is a peptide derived from the ghrelin precursor, includes 23 amino acids, and has an opposite function to ghrelin. It is associated with decreased gastrointestinal motility, reduced appetite and food intake, and weight loss.¹⁴ Although limited studies have investigated the relationship between obestatin and IBDs, none have found a significant correlation between obestatin levels and disease activity.^14,15 However, 2 performed studies by Alexandridis et al¹⁴ and Jung et al¹⁵ demonstrated a decrease in the obestatin/ghrelin ratio, which may indicate disease activity. However, the finding of Kim et al’s¹⁶ study did not support this conclusion.

In this study, we conducted a systematic review and meta-analysis with the aim of shedding light on the relationship between peripheral levels of ghrelin and obestatin with IBDs. Additionally, we examined the impact of infliximab on patients’ ghrelin levels and explored the relation between circulating ghrelin levels and bone mineral density in IBD patients.

Methods

Search Strategy and Study Selection

Our systematic review and meta-analysis were done according to the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) 2020 statement.¹⁷ The searches were conducted in PubMed, Embase, Scopus, and the Web of Science on May 13, 2023, and all articles until that point were included. The main search terms were “inflammatory bowel disease*,” “Crohn*,” “Crohn’s disease,” “ulcerative colitis,” “ghrelin,” “hunger hormone,” and “obestatin.” The detailed search queries can be found in Supplementary Table 1. This systematic review’s protocol was registered in PROSPERO (CRD42023432126).

Two independent reviewers (A.K. and A.H.B.) screened all the studies, first by title and abstract, and then by reviewing the full text. A third reviewer (N.B.) resolved the disagreements and finalized the included studies. The included studies had to measure the plasma and/or serum level of ghrelin and/or obestatin in patients with active or remitted IBDs (UC or CD) compared with healthy control subjects. Conference abstracts, protocols, or review articles were all excluded. Studies that evaluated these markers in tissues, in vitro studies, and in vivo studies were also excluded. The complete process of identifying and selecting the studies is summarized in Figure 1.

Data Extraction

The data extraction of the included studies was done using Microsoft Excel spreadsheet software. One reviewer extracted the prespecified data (A.K.) and a second reviewer (A.H.B.) cross-checked the extracted data. The following data were extracted from the included studies: (1) first author name, publication year, country of publication, and study design; (2) study population and type of IBD (UC or CD); (3) number of participants in each group, mean age ± SD, body mass index (BMI), disease duration, and male percentage in total population; (4) plasma or serum concentration of ghrelin and obestatin; and (5) main findings of the articles.

Quality Assessment

We assessed the methodological quality of the studies by using the checklist known as the Newcastle–Ottawa Scale (NOS)¹⁸ by assigning 2 reviewers to evaluate them (A.K. and A.H.B.). According to the NOS, selection, comparability, and outcome were assessed as potential biases. Each of these parameters was categorized as very good (score of 9-10), good (score of 7-8), satisfactory (score of 5-6), or unsatisfactory (score of 0-5).

Statistical Analysis

All our analyses were done using STATA (version 17.0; StataCorp). We used a random-effects (restricted maximum likelihood) model for the meta-analyses. The analyses were done for (1) ghrelin in patients with active IBD, UC, and CD, compared with ghrelin in patients who had remission; (2) ghrelin in patients with active IBD, UC, and CD, compared with healthy control subjects; and (3) obestatin level and obestatin-to-ghrelin ratios in patients with active IBD, UC, and CD, compared with these values in patients with the remitted disease. Also, we performed a leave-one-out sensitivity analysis for the meta-analysis of ghrelin levels in patients with active IBD vs quiescent IBD and for the meta-analysis of ghrelin levels in patients with active IBD vs healthy control subjects.

We calculated the standardized mean difference (SMD) and 95% confidence interval (CI) for comparison of the biomarkers (ghrelin or obestatin) levels in IBD patients and control subjects. SMD is used in analyses with different settings and methods of measurement and is considered better than the mean difference when different specimens (serum/plasma) with different kits are used.¹⁹ In some studies, the level of the biomarkers was reported in the median and interquartile range (IQR) or median and range. We used the methodological ways suggested by Luo et al²⁰ and Wan et al²¹ to convert them into mean and SD. Subgroup analysis based on serum/plasma specimens was also performed. The heterogeneity of studies was calculated using Cochrane’s Q statistic²² and Higgin’s I² and H² tests.²³ We considered heterogeneity thresholds as ≤25% for low, 26% to 75% for moderate, and >75% for high. Publication bias was assessed by visual inspection of funnel plots in addition to Begg’s and Egger’s tests.^24,25 Finally, for comparison of active vs quiescent IBD, we performed meta-regression based on publication year, mean age, mean BMI, and sample size. The related bubble plots were also designed.

Results

Literature Search and Included Studies

The initial search yielded 752 results, including 119 from PubMed, 181 from Embase, 185 from the Web of Science, and 267 from Scopus. Duplicates were removed (n = 297) and the remaining 455 studies were screened based on titles and abstracts. At this stage, 37 studies remained and went through full-text screening. After exclusion for various reasons, 19 studies were included in this review.^7-16,26-34 The selection process and exclusion reasons are described in Figure 1.

Table 1 shows the details of included studies. Studies were published between 2006 and 2023. Greece was the country with 3 included studies,^7,13,14 followed by Turkey,^8,26 South Korea,^15,16 China,^27,34 and the United Kingdom,^28,32 each having 2 studies. Totally, 1064 patients with IBD (476 UC and 588 CD) and 388 healthy control subjects were studied. The qualities of all included studies were high based on the NOS quality assessment method (Table 2).

Meta-Analysis of Ghrelin Levels in Patients With Active IBD vs Patients in Remission

Eleven studies were included that compared ghrelin levels between active IBD and those in remission.^{8-11,14-16,26,27,29,30} Meta-analysis of these studies showed that active patients had significantly higher levels of ghrelin (SMD, 0.70; 95% CI, 0.06 to 1.34; P = .03) (Figure 2), while the heterogeneity was high for this analysis (I² = 91.7%). In subgroup analysis based on serum/plasma specimens, however, there was no significant difference between groups (P = .63). Also, these 2 subgroups showed no difference between active IBD and remission (serum: SMD, 0.81; 95% CI, −0.05 to 1.68; P = .07; plasma: SMD, 0.41; 95% CI, −0.09 to 0.91; P = .11).

Analyzing 4 studies assessing UC patients only showed no significant difference (SMD, 1.30; 95% CI, −0.28 to 2.88; P = .11) (Figure 3A). Moreover, a similar result was obtained in studies evaluating patients with CD only (SMD, 0.80; 95% CI, −0.41 to 2.01; P = .20) (Figure 3B).

Publication bias was investigated for the comparison of active and quiescent IBD, and there was no asymmetry in the contour-enhanced funnel plot (Supplementary Figure 1). In a similar way, neither Begg’s nor Egger’s test could reveal any source of publication bias (P = .75 and .53, respectively). Leave-one-out sensitivity analysis of this analysis, however, showed that by each removal of 5 of the studies,^8,9,14,15,26 the overall difference becomes insignificant (Supplementary Figure 2).

Meta-regression using the variables including sample size, publication year, mean age, and mean BMI was performed, the results of which are shown in Supplementary Table 2. It was shown that publication year had a significant association with the pooled effect size (P = .047). Moreover, it accounted for 27.83% of the variance. The funnel plots for these analyses are shown in Supplementary Figures 3-6.

Meta-Analysis of Ghrelin Levels in Patients With Active IBD vs Healthy Control Subjects

Eight studies compared the ghrelin levels in patients with active IBD and healthy control subjects.^{8-11,27,29,30,34} The pooled estimate through meta-analysis of these studies showed that there was no significant difference between the groups (SMD, 0.44; 95% CI, −0.39 to 1.27; P = .30) (Figure 4). However, the heterogeneity was high (I²: 94.2%).

Separate analysis for UC and CD patients showed that none of these 2 subgroups of IBD had different ghrelin levels (SMD, 0.37; 95% CI, −1.36 to 2.10; P = .68; and SMD, 1.20; 95% CI, −0.43 to 2.83; P = .15, respectively) (Figure 5A and 5B respectively).

The funnel plot did not reveal any source of asymmetry (Supplementary Figure 7). Also, Begg’s and Egger’s tests did not reveal any significant bias (P = .39 and .10, respectively). Leave-one-out by sensitivity analysis method resulted in no difference from any studies’ omission (Supplementary Figure 8).

Meta-Analysis of Ghrelin Levels in Patients in Remission vs Healthy Control Subjects

A meta-analysis of patients with IBD and in remission in comparison with control subjects showed that there was no statistical difference between the groups in terms of ghrelin levels (SMD, −1.47; 95% CI, −3.29 to 0.35; P = .11) (Figure 6). This analysis had a high heterogeneity (I² = 98.6%).

The funnel plot for this analysis did not reveal any sign of asymmetry and publication bias, as shown in Supplementary Figure 9. Egger’s and Begg’s statistical tests, however, showed significant publication bias, either (P < .001 and P = .072, respectively). The leave-one-out method sensitivity analysis also revealed a significant change in pooled results by the removal of the Nishi et al¹⁰ study, representing an SMD of −0.51 (95% CI, −0.98 to −0.03; P = .036) (Supplementary Figure 10).

Meta-Analysis of Obestatin Levels and Obestatin/Ghrelin Ratio in Patients With Active IBD vs Patients in Remission

Obestatin levels were analyzed by a meta-analysis of 3 studies,^14-16 and it was shown that there was no significant difference between the groups (SMD, 0.31; 95% CI, −0.05 to 0.68; P = .09) (Figure 7A). However, the analysis of the obestatin/ghrelin ratio resulted in a significantly lower ratio in patients with active IBD, compared with control subjects (SMD, −1.90; 95% CI, −2.45 to −1.35; P < .01) (Figure 7B).

Discriminatory Power of Ghrelin in Patients With IBD

Three studies evaluated the discriminatory ability of ghrelin in IBD patients.^26,27,29 Cekic et al²⁶ found that ghrelin can distinguish active IBD from those in remission with excellent performance (area under the receiver-operating characteristic curve [AUC], 0.934; 95% CI, 0.884-0.984; P < .001). In another study evaluating UC patients, Lv et al²⁷ found an AUC of 0.788 (95% CI, 0.690-0.894) with a susceptibility of 82.35% and a specificity of 90.00% in distinguishing active UC from those in remission (P < .001). Ortega Moreno et al²⁹ evaluated the discriminatory ability of ghrelin in CD patients and found AUCs for discriminating active CD from quiescent CD (AUC, 0.64; 95% CI, 0.46-0.82), active CD from healthy control subjects (AUC, 0.68; 95% CI, 0.52-0.84), and quiescent CD from healthy control subjects (AUC, 0.54; 95% CI, 0.38-0.69). These 3 AUCs were pooled using random-effects meta-analysis, and the overall AUC of 0.81 (95% CI, 0.65-0.96) was found (Supplementary Figure 11).

Association Between Ghrelin and Osteoporosis in IBD Patients

Koutroubakis et al¹³ evaluated patients with IBD and compared ghrelin levels between normal (n = 40 [33.9%]), osteopenic (n = 55 [46.6%]), and osteoporotic (n = 23 [19.5%]) patients. IBD patients with osteoporosis had significantly higher levels of ghrelin compared with the normal and osteoporotic groups (P = .038).

Ghrelin Levels and Infliximab Therapy in CD Patients

Two studies evaluated the effect of infliximab therapy on ghrelin levels in CD patients.^12,32 Lönnkvist et al¹² evaluated change in ghrelin levels after infliximab infusions in CD patients. They found decreased ghrelin after infliximab (initial infusion: from 786 [IQR, 649-1160] pg/mL to 591 pg/mL [IQR, 508-817]; P < .05; 6-month follow-up infusion: from 784.5 [IQR, 459.3-1185] pg/mL to 635.5 [IQR, 509-874.5] pg/mL; P = .06). It should be noted that these values were not different from control subjects (729.5 [IQR, 605-823.8] pg/mL; P > .05). However, a study conducted by Sung et al³² found that 2-hour integrated total ghrelin significantly increased after infliximab therapy in 11 patients with CD (from 162 [95% CI, 99-311] pg/mL.h to 200 [95% CI, 128-387] pg/mL.h; P = .02).

Discussion

IBDs encompass UC, CD, and indeterminate colitis. These subtypes differ in terms of symptoms, pathology, therapeutic outcomes, rate of remission, and exacerbation. The conventional method of diagnosing IBD involves endoscopic procedures and biopsies. Confirmatory diagnosis of these subtypes is also challenging, highlighting the need for a noninvasive and available biomarker. There is still no single specific biomarker for predicting treatment response and monitoring the disease progression in patients with IBD. In our study, we compared levels of ghrelin, obestatin, and obestatin/ghrelin ratio in patients with active IBD, those patients in remission, and non-IBD control subjects. Our main findings revealed significantly higher ghrelin concentration in patients with active IBD compared with those with quiescent IBD, as well as a lower obestatin/ghrelin ratio in patients with active IBD in comparison with patients in remission. However, the lack of significant difference between active/quiescent IBD and healthy control subjects shows that this might not be the ideal biomarker for this comparison. Hence, the search for a promising biomarker should be continued.

Ghrelin is an endogenous hormone primarily secreted by the neuroendocrine cells of the stomach, and it is also secreted to a lesser extent by small and large intestines.^35-37 Ghrelin exerts various physiological effects: it is a stimulator of growth hormone and an antagonist of leptin, which plays a role in controlling food intake, causing weight gain, and decreasing fat utilization.^38,39 Studies have shown that ghrelin concentrations decrease in individuals with high BMI and obesity can blunt ghrelin’s effect.^36,40-42 Ghrelin is involved in gastrointestinal tract activity, and by increasing intestinal movement it may have a role in the recovery of patients with IBD.⁴³ It may also benefit gastrointestinal mucosal stability through increased colon blood flow and promotion of epithelial cell proliferation.⁴⁴ Additionally, ghrelin exhibits anti-inflammatory effects on the gastrointestinal tract by inhibiting proinflammatory cytokines such as IL-1β, IL-6, and TNF-α.⁴⁵ It can modulate immune responses and contribute to the control of inflammation through the immunoregulation of T cells and macrophages.⁴⁵

Obestatin is a recently discovered hormone coexpressed with ghrelin in X/A-like cells of the gastric fundus. Derived from preproghrelin, obestatin is known to decrease food intake, inhibit weight gain, and reduce jejunal motility.⁴⁶ However, there are conflicting results regarding its potential anti-inflammatory effects, and further research is needed to fully understand this aspect.

Some studies evaluated tissue ghrelin in IBD patients.^47,48 Sahin et al⁴⁷ compared tissue ghrelin between UC patients and healthy control subjects. They found no difference between patients and control subjects in the amount of staining with ghrelin; however, colonic ghrelin staining was associated with higher disease activity in UC patients. However, they have not measured tissue ghrelin messenger RNA concentration in samples from patients and control subjects. Another study by Hosomi et al⁴⁸ found significantly higher mucosal messenger RNA levels of ghrelin in patients with IBD (both UC and CD) in comparison with healthy control subjects. They suggested that ghrelin may contribute to CD pathogenesis by reactivity dysregulation in peripheral T cells that may lead to T helper 1–skewed inflammation.

Our study revealed a significant difference in ghrelin levels between patients with active IBD and those with quiescent IBD, consistent with previous reports.^8,9,14,49 However, we did not find a significant difference in ghrelin levels when comparing patients with active or quiescent UC. Similarly, no difference in ghrelin levels was observed between patients with active or quiescent CD. This discrepancy may be attributed to the fact that some of the included studies reported only the total number of patients with IBD without classifying them based on specific subtypes. Furthermore, our data revealed no significant difference in ghrelin levels between patients with active IBD and the control group. The same result indicates for patients with CD or UC when compared with their control group.

Strengths and Limitations

Although in this study the obestatin levels did not differ between patients with active IBD and remission, the obestatin/ghrelin ratio was significantly lower in patients with active IBD. However, it is important to interpret this result with caution considering the limited number of articles investigating obestatin levels. This meta-analysis has some limitations. First, a small number of patients were included in each study, making it difficult to apply the result to other populations. Second, the heterogeneity of BMI between patients with IBD and the control group in Ghomraoui et al⁴⁹ was considerable and might affect the outcome. Third, the duration of the disease has not been mentioned in some of the included articles, whereas several studies have indicated a potential relationship between ghrelin levels and the duration of IBD.¹³ Fourth, ghrelin levels are known to vary in response to many clinical conditions including circadian rhythm, fasting and postprandial timing, and nutrient composition such as protein, fatty acid, or glucose.⁵⁰ Because these conditions were not routinely controlled across the reported studies, they may contribute to variations in measured values. Fifth, the high variation in values for normal individuals suggests that some of the assays may be more accurately measuring the target ghrelin, while others may be missing or measuring artifacts. Finally, due to a lack of data regarding the type of ghrelin (acyl and des-acyl ghrelin) used in the included studies, we were not able to perform subgroup analysis based on the ghrelin type.⁵¹ While this study provides insight into the differences in mentioned biomarkers, further research with a larger sample size and considering disease duration and therapies are needed for identifying potential biomarkers. Moreover, such studies should utilize validated standards and controlled clinical conditions for blood sampling and processing.

Conclusion

To the best of our knowledge, the current systematic review and meta-analysis is the first one that compares ghrelin, obestatin, and obestatin/ghrelin ratio in IBDs and healthy control subjects. We found significantly higher ghrelin levels in active IBD patients in comparison with those in remission in addition to a lower obestatin/ghrelin ratio in IBD patients when compared with control subjects. We believe that our study could pave the way for future research to investigate appropriate adipokines as biomarkers of IBD or active IBD. More studies with higher sample sizes evaluating the specificity and sensitivity of these markers are warranted to confirm our findings.

Author Contribution

A.K.: Writing—original draft, review & editing/Conceptualization/Formal analysis/Visualization. N.B., Z.A., B.S.: Writing—original draft/Data curation. S.P., N.A.B.: Writing—review & editing. A.H.B.: Supervision/Writing—review & editing. All authors read and approved the final manuscript.

Funding

None received.

Competing interests

The authors declare that they have no competing interests.

Consent for publication

Not applicable

References