Gut microbial balance and liver transplantation: alteration, management, and prediction

Abstract

Liver transplantation is a conventional treatment for terminal stage liver diseases. However, several complications still hinder the survival rate. Intestinal barrier destruction is widely observed among patients receiving liver transplant and suffering from ischemia–reperfusion or rejection injuries because of the relationship between the intestine and the liver, both in anatomy and function. Importantly, the resulting alteration of gut microbiota aggravates graft dysfunctions during the process. This article reviews the research progress for gut microbial alterations and liver transplantation. Especially, this work also evaluates research on the management of gut microbial alteration and the prediction of possible injuries utilizing microbial alteration during liver transplantation. In addition, we propose possible directions for research on gut microbial alteration during liver transplantation and offer a hypothesis on the utilization of microbial alteration in liver transplantation. The aim is not only to predict perioperative injuries but also to function as a method of treatment or even inhibit the rejection of liver transplantation.

Keywords

gut microbial balance / liver transplantation / ischemia–reperfusion / acute rejection

Introduction

During the last few decades, orthotopic liver transplantation (OLT) has been used as conventional therapy for nonreversible acute and chronic liver diseases [¹,²]. The number of patients undergoing OLT has increased dramatically, and the success rate of such operations has improved steadily [³]. However, perioperative injuries caused by ischemia–reperfusion (I/R) or acute rejection (AR) still hinder the survival rate.

Currently, intestinal microbiota is identified as an extremely important microecosystem and is even considered as a main metabolic “organ” that maintains a symbiotic relationship with the body [⁴–⁶]. Between 10 and 100 trillion microbes are considered to live in the human gastrointestinal tract [⁷,⁸]. Their importance in resisting pathogenic microbes [⁹–¹¹], regulating metabolic processes [⁸,¹²], functioning as a neglected endocrine organ [¹³], and inducing the maturation of the immune system, have been demonstrated [¹⁴]. Nevertheless, intestinal microbiota has been related to a variety of diseases, such as inflammatory bowel disease [¹⁵,¹⁶], hepatosis, cardiovascular diseases [¹⁷], obesity [¹⁸], diabetes [¹⁹,²⁰], colorectal cancer [²¹,²²], and nervous system diseases [²³].

The status of health or disease is related to the closely linked anatomies and functions of the intestine and liver. According to current findings, the majority of intestinal microbiota (autochthonous groups, such as the Bifidobacterium and the Bacteroides fragilis subgroup) of a healthy host presents host-specific relative stability over time within each individual [²⁴], whereas the total bacteria of gut microbiota (such as Faecalibacterium prausnitzii and Bifidobacterium spp.) changed dramatically in cirrhotic patients [²⁵]. In a study that assessed the fecal Lactobacilli population, low diversity index was found in patients with hepatitis B virus (HBV)-related decompensated cirrhosis [²⁶].

During liver transplantation, the gut barrier can be ravaged, and gut microbiota can be impeded in hepatic I/R or AR injuries [²⁷,²⁸]. Thus, significant alteration of abundant intestinal bacteria exists causing severe infection [²⁹]. Therefore, the appropriate control and prevention of specific infections instituting a gut microbial balance should be applied. In addition, the possibility of utilizing microbiota as a biomarker in predicting perioperative injuries of OLT should be discussed.

Alteration

I/R injury

Xing et al. [²⁷] found a dramatic alteration of microflora in model rats with hepatic I/R injury. In the hepatic I/R group, serum alanine aminotransferase and aspartate aminotransferase levels apparently increased with the elevation of malondialdehyde concentration and reduced superoxide dismutase activity. In addition, counts of Bifidobacteria and Lactobacillium, two anaerobic bacteria, were found to be low. Nevertheless, the counts of Enterobacteriaceae and Enterococci were high. Such alteration was found to have a close correlation with the destruction of epithelial apical surface, damage of ileal epithelial tight junction, increase in plasma endotoxin, and a high possibility of bacterial transplantation to the kidney. Thus, we concluded that liver I/R injury can lead to the alteration of microflora and destruction of the gut mucosal barrier function, which may result in bacterial translocation and endotoxemia.

AR injury

Despite the wide use of OLT, AR remains a life-threatening complication even today. The interactions between gut microbiota and the host in AR injury after OLT still need to be explored. A study was designed to monitor microbial alteration in rats with AR after OLT utilizing polymerase chain reaction-denaturing gradient gel electrophoresis (PCRDGGE) and reverse transcription quantitative real-time polymerase chain reaction. Interestingly, the study found that in rats with AR, gut microbial alterations were related to an increase in plasma endotoxin and an increased possibility of bacterial translocation. Moreover, among the six major microorganisms that were studied, the excessive growth of Bacteroides and Ruminococcus significantly dominated the gut microbial structure shifts [²⁸].

Damage to the intestinal barrier

Under physiological conditions, the gastrointestinal system not only digests and absorbs foods but also functions as a defensive barrier [³⁰]. However, intestinal barrier dysfunction can be widely seen not only in the rats with OLT bearing I/R injury or AR injury but also in rats with preoperative malnutrition, which is very common in chronic hepatic disease patients [³¹,³²]. Different degrees of intestinal barrier damage can be seen as follows: motility weakening, structure impairment, and the secretion reduction that may damage the physical barrier [³³]. Moreover, microbial dysbiosis and juiceless digestive secretion damage the chemical and biological barriers [³⁴]. Shrinkage of the intestine-related lymphoid tissue and a decrease in the gut secretion of sIgA abate the immune barrier [³⁰]. Thus, protection of the intestinal barrier utilizing microbial balance is important.

Management

Ameliorating I/R injury

A significant number of studies have shown the possible mechanism of I/R injury [³⁵]. However, only a handful of studies have concentrated on the vital role of intestinal microbiota.

A study showed that the use of probiotics, such as Bifidobacterium catenulatum ZYB0401 plus Lactobacillus fermentum ZYL0401, can prevent hepatic I/R injury in rats by modifying the gut microflora, ameliorating the intestinal mucous barrier, suppressing the possibility of bacterial transplantation and plasma endotoxin levels, and ameliorating hepatic lipid peroxidation. Thus, the author recommended a combined utilization of different varieties of probiotics [¹⁶].

Liver ischemic preconditioning (IPC) ameliorates I/R injury after OLT. In a recent study conducted by Ren et al. [³⁶], they revealed a protective effect of liver IPC by the restoration of intestinal microbiota and the improvement of gut barrier function following OLT in rats (Fig. 1). Moreover, intestinal microbial diversity and the richness of the species increased. Furthermore, in liver IPC rats, they identified ten key classes of microbic structural shifts, the majority of which were classified to phylum Bacteroidetes. Overall, these changes were beneficial for the balance and stability of gut microflora, indicating the beneficial effect of liver IPC in OLT. Therefore, the author speculated that the function of a hepatic graft can be enhanced by liver IPC during liver transplantation through the restoration of the gut microbial balance and even positive feedback of the “gut-liver axis.”

Reducing AR injury

Although an alteration of the intestinal microbiota was observed in rats with AR injury after OLT, few studies have been dedicated to the mechanisms or the treatment of AR injury utilizing microbial balance. Recently, Xie et al. [³⁷] compared the effect of probiotics and antibiotics on hepatic damage in rats with AR injury after OLT. Several parameters, including gut microbial alterations, hepatic histopathology and function, cytokines, and T cells, were observed. Although the liver injury was remarkably ameliorated either by treatment with antibiotics or probiotics, significant isolated diversity of fecal bacteria was seen in the probiotic group. Importantly, with probiotic treatment, the diversity of gut microbiota and the pathology of the ileum and colon of the AR rats showed dramatic improvement compared with the antibiotic and the control groups, especially regarding the amounts of Lactobacillus and Bifidobacterium. Moreover, the author noted the differences in Treg cells, tumor growth factor (TGF)-b and CD4/CD8 between the three groups and concluded that, although both antibiotics and probiotics can ameliorate hepatic injury in rats with AR after OLT, different mechanisms were involved and probiotics were the best option. In addition, the study suggested that probiotics prevent AR injury through augmentation of Treg cells and TGF-b and the amelioration of CD4/CD8. This finding may become a potential focus for research on the protective mechanisms of probiotics on AR injury.

Strengthening intestinal barrier

Several studies were published recently on the protection of the intestinal barrier in malnourished rats after OLT. Ren et al. [³²] reported that the supplementation of probiotics can reduce the level of serum endotoxin and the opportunities for bacterial translocation. This treatment also promoted the recovery of intestinal barrier functions in a study of malnourished rats after liver transplantation. Furthermore, decreased concentrations of serum tumor necrosis factor (TNF)-α and attenuated inflammatory cascade reactions in the probiotics group also suggested a mild inflammatory response induced by the OLT operation. Furthermore, this conclusion regarding the protective effect of probiotics on immunomodulation and health was obtained by the increased intestinal secretion of sIgA, promoted proliferation of lymphocytes in intestinal Peyer’s patches, and enhanced alteration of lymphocyte phenotypes in intestinal Peyer’s patches.

Furthermore, the mucosal ultrastructure observed by Jiang et al. [³¹] indicated improvement of intestinal protein metabolism following the protection of intestinal mechanical barrier with supplementation of glycyl-glutamine in the rats after OLT. Six clusters of aerobic bacteria, namely, Escherichia coli, Enterococcus, Proteus vulgaris, Streptococcus agalactiae, Proteus mirabilis, and Viridans streptococci, were noted to display significant decreases in counts with the supplementation of glycyl-glutamine. The decrease in plasma endotoxin and TNF-α also suggested a protective effect. Moreover, increased ileocecal sIgA in the glycyl-glutamine group improved the immune barrier of the intestines. Thus, the author provided another method for the protection of intestinal barrier by the supplementation of glycyl-glutamine.

Prediction

In the past, only 10%–30% of gut microbiota can be detected by classical culturing techniques [³⁸]. In recent years, progressive molecular technologies, such as next-generation sequencing technologies, including 16S rRNA sequencing [³⁹] and metagenomic sequencing [⁴⁰], and PCRDGGE has benefited the measurement of intestinal microbiota and the sensibility of predicting perioperative injuries.

With the application of natural-source glycogen measuring lactobacilli 16S rDNA, extracted from fecal samples, Wu et al. [²⁶] were able to find that the Lactobacillus floras in either the group of patients with HBV-related decompensated cirrhosis or the group with liver transplants for hepatitis B cirrhosis were significantly different from that in healthy subjects. Compared with healthy people, decreased diversity of fecal lactobacilli was found in the patients, which was prominent in the group of decompensated cirrhosis. This finding may provide a method of predicting the cirrhotic patients who need to receive OLT.

Similarly, to identify the intestinal bacterial populations, a retrospective analysis conducted by Lu et al. [²⁹] used several data from digitized denaturing gradient gel electrophoresis fingerprints. Compared with normal individuals, the diversity of the prominent intestinal microbiome of OLT patients reduced dramatically. Decreases in all Eubacteria, Bifidobacterium spp., F. prausnitzii, and Lactobacillus spp. can be easily seen in liver recipients. The destruction of the microbiota, which is attributed to the empiric prophylaxis strategy, may increase the vulnerability of patients after OLT and increase opportunistic pathogens, such as Bifidobacterium dentium. Thus, cognizance of the intestinal microbial profile may predict injuries caused by the abuse of antibiotics.

Analyzing the peculiarity of the ileal microbiota in small bowel transplantation, Oh et al. [⁴¹] stated that microbial profiling may have the potential to become a biomarker in the diagnosis of small bowel transplantation rejection. In a pilot study of kidney transplantation, AR was associated with an increase in the relative abundance of Proteobacteria and a decrease in Firmicutes [⁴²]. In addition, a study of stem cell transplant recipients revealed that the graft-versus-host disease can be predicted by the measurement of microbial alterations [⁴³]. Recently, Ren et al. [⁴⁴] analyzed different parameters in rats with OLT, indicating that the gut microbial variation preceded barrier dysfunction and that the intestinal microbial alteration was more sensitive than hepatic histology in predicting hepatic acute or chronic rejection injury after OLT (Fig. 2). Furthermore, four key bands of microbial variation in AR injuries were explored, providing a feasible auxiliary biomarker for the diagnosis of early AR after OLT.

Conclusions

As a result of the progress in microbial detection technologies, although perioperative injuries especially I/R, AR or subsequent intestinal barrier dysfunctions restrict the use of OLT, accurate mechanisms and the management of reducing injury through microbial balance are rapidly being revealed. Furthermore, gut microbial profiles as perioperative biomarkers for injury are showing potential in injury prediction.

Perspectives

Although a variety of protection methods, including antibiotics, probiotics, and prebiotics, are being used today, definitive dosages and the use of the treatment remain unclear. The use of probiotics or intestinal decontamination during the perioperative period is still especially controversial today. Thus, more studies revealing exact mechanisms are strongly desired.

Probiotics have been shown to prevent AR injury of liver transplantation. Moreover, the common clinical immunosuppressors, such as CsA and FK 506, are all products of the metabolites of the microbiota. Thus, microbial metabolites from patients with immune tolerance should be studied further to determine whether the product of the microbiota can induce immune tolerance after liver transplantation.

Nevertheless, as one of the key methods for diagnosis in the future, the veracity and the quantity of microbial biomarkers should be studied extensively. In addition, studies should concentrate on whether microbial alteration is the cause or part of the sequence of injury after OLT.

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