Effects of synbiotic supplementation on energy and macronutrients homeostasis and muscle wasting of critical care patients: study protocol and a review of previous studies

The gut microbiota refers to the commensal microorganisms that reside in our gastrointestinal tract (GIT) with a symbiotic relationship [1]. Gut microbiota has a significant role in host metabolism and homeostasis [2, 3]. A disturbance in this microbial community, which leads to an unhealthy state, is called dysbiosis [4]. Over the past decade, emerging evidence has demonstrated the role of intestinal dysbiosis in the pathogenesis of various conditions, such as infectious, immune, and metabolic diseases [5], while it has not been studied extensively in critical illness. An extreme and persistent dysbiosis occurs among critically ill patients, regardless of the heterogeneity of disease. The extreme dysbiosis in patients under critical care is due to the stress of critical illness, multiple antibiotics and additional pharmacological interventions, and highly processed enteral/parenteral nutrition [6, 7]. Dysbiosis in critically ill patients may make them prone to hospital-acquired infections, sepsis, multi-organ failure (MOF), energy homeostasis disturbance, muscle wasting, and cachexia [6, 8, 9].

The majority of patients in the intensive care unit (ICU) have had a severe illness, trauma, or major surgery, and accordingly they are unable to manage their nutritional demands. Although nutritional support is a daily practice in the ICU, many patients still suffer from malnutrition due to lack of intake or uptake of nutrients [10]. The prevalence of malnutrition in the ICU within developed and developing countries is reported as 50.8% and 78.1%, respectively [11]. Malnutrition is independently associated with longer ICU stay, more ICU readmissions, and a higher incidence of infections and risk of mortality [11]. A greater degree of malnutrition is also associated with a higher risk of 28-day mortality [12]. Malnutrition further tends toward acute or chronic loss of muscle bulk and function [13]. The gut microbiota and their derived metabolites play an essential role in the absorption, storage, and consumption of energy derived from the diet [14, 15]. Previous studies suggest that modulating gut microbiota by novel therapeutics, such as prebiotics, probiotics, or synbiotics, can have an effect on gastrointestinal tolerance and complications of enteral nutrition, which eventually lead to the regulation of energy intake. Recently, Tuncay et al. showed that enteric formula with prebiotic content in patients under neurocritical care was associated with a significant increase in total feed volume and energy intake and a non-significant tendency to achieve a target dose of nutrition more frequently and earlier [16]. Malik et al. also found that in patients in the ICU, receiving enteral formula supplemented with probiotics led to a faster return of gut function (tolerated feeding of 80% of their estimated required calories for 48 h consecutively) [17]. However, Sanaie et al. demonstrated that daily energy and protein intake in patients receiving probiotic supplements on the ICU were not significantly different from the group receiving placebo [18]. In the critical care setting, diarrhea is the most obvious complication of enteral nutrition (EN), which is associated with the inadequacy of energy and macronutrient intake [16]. Previous systematic reviews have confirmed the significant benefit of probiotics in the reduction of diarrhea in hospital patients overall. But a recent meta-analysis focused on patients in the ICU found no benefit of probiotics in preventing or treating diarrhea [19]. Besides, in the dysbiosis state of critical illness, microbial products that reach distant organs like brain, adipose tissue, and liver, favor the development of immune-mediated diseases and metabolic alterations [20]. Gut microbiota metabolites like short-chain fatty acids (SCFAs) can also influence the immune system and host metabolism, which regulates energy homeostasis [20]. Thus, gut microbiota modulation may be beneficial in the regulation of immune and metabolic responses and energy homeostasis.

Muscle wasting, characterized by loss of muscle mass and strength, is associated with negative health outcomes such as functional disability, greater risk of infections, delayed recovery, poor life quality, and mortality [21]. The gut microbiota have been reported to influence muscle metabolism. The molecular mechanisms of this gut-muscle axis remain to be identified. Gut microbiota influence amino acid bioavailability and are sources of different metabolites, such as conjugated linoleic acid, acetate, and bile acids, which modulate muscle metabolism [8]. Various pathogen-associated molecular patterns (PAMPs) activate the transcription factor nuclear factor kappa light chain enhancer of activated B cells (NF-kB) through toll-like receptors (TLRs) and modulate the production of proinflammatory cytokines, which can induce muscle atrophy [8] (Fig. 1).

Modulation of gut microbiota through prebiotics, probiotics, or synbiotics can be considered as a potential treatment for muscle wasting and cachexia. In mouse models of leukemia, restoring Lactobacillus species by oral supplementation with Lactobacillus reuteri 100–23 and Lactobacillus gasseri 311,476 reduces inflammatory cytokines and expression of muscle atrophy makers [22]. In another study, Bindels et al. showed that prebiotic supplementation in leukemic mice could contribute to delaying anorexia and fat mass sparing by inducing a metabolic shift in adipose tissue [23]. In the mouse models of cancer cachexia, administration of a synbiotic supplement including inulin-type fructans and live L. reuteri 100–23 was associated with restoration of the gut barrier and immune function, thus reducing cachexia. It also prolonged survival [24]. Varian et al. also showed that probiotic administration in leukemic mice could inhibit cachexia by reducing systemic inflammation [25].

Considering the extreme dysbiosis in critically ill patients and related energy and macronutrients homeostasis disturbance and muscle wasting, prompted us to evaluate the effect of synbiotic supplementation on the elimination of this condition. To our knowledge, this is the first study to investiage the effect of synbiotic supplementation on muscle wasting in critically ill patients.

The primary objective is to evaluate the effects of synbiotic supplementation on energy and macronutrient homeostasis and muscle wasting in patients under critical care.

The secondary objective is to evaluate the effects of synbiotic supplementation on infectious complications and length of hospital and ICU stay in patients under critical care.

Participants must meet all the inclusion criteria to participate in this study: adults aged 18–65 years; admitted to the ICU; hemodynamically stable within 24–48 h after admission; requiring exclusive enteral nutrition (EN) via a nasogastric tube (NGT); not taking any kind of microbial cell preparations (prebiotics probiotics, synbiotics); estimated length of ICU stay more than 14 days; and provision of written consent.

All candidates meeting any of the exclusion criteria at baseline will be excluded from study participation: pregnancy or lactation; any contraindication to EN; any contraindication to insertion of the NGT; receiving immunosuppressive treatment, radiotherapy, or chemotherapy; hematologic disease; acquired immune deficiency syndrome (AIDS); transplant recipient; known allergy to microbial cell preparations; cancer or autoimmune diseases; artificial heart valve or congenital heart valve disease; estimated length of ICU stay less than 4 days; gastric disease; or gastrointestinal tract (GI) tract surgery.

Before the screening procedure, informed consent will be obtained from every participant who meets the inclusion criteria. First, we will describe the purpose of the study, the procedures involved, the length of time the subject is suspected to participate, any possible disadvantages or discomforts, the benefits of the study to society and individuals, and the person to contact for answers to further questions. We will also emphasize that participation is voluntary, and refusal or withdrawal will not cause any loss of benefits that they are entitled to receive. Then the participants or their legal guardian will read and sign two copies of the written form. If, the informed consent was obtained from the patient’s guardian because of the patient’s lack of competence to consent and then later in the study the patient subsequently became competent as required, consent will be regained.

After providing their written consent, patients are randomly allocated in a 1:1 ratio to the intervention or control group (A or B). Patients will be randomized through a stratified sequential randomization plan generated online. Randomization will be stratified by disease severity (Acute Physiology and Chronic Health Evaluation (APACHE) II,¹ 0–35 and 35–70). For allocation concealment, we will use sealed opaque envelopes; inside each there is a carbon paper and the A or B card. To avoid probable selection bias, we will write the patient’s name on the envelope before opening it. All patients, researchers, and medical staff will be blinded to the allocation to either synbiotic or placebo capsules. An available third party, the secretary of the ICU, will be aware of whether A or B is allocated the synbiotic supplement. In the case of any complication associated with the intervention allocated, the medical staff will refer to the secretary for details.

All eligible patients will receive standard hospital gavage as EN through the NGT in the 24-48 h after admission to the ICU. According to the recent European Society of Parenteral and Enteral Nutrition (ESPEN) guideline on clinical nutrition in the ICU [26], continuous rather than bolus EN is preferred because it causes less diarrhea, but there is no difference in other outcomes. Another systematic review showed that bolus feeding is associated with lower aspiration rate and better calorie attainment [27]. It also provides a greater stimulus for protein synthesis [28]. Considering these data and the availability of bolus EN in our hospitals we applied this method. In the absence of an indirect calorimeter, the simple weight-based equation of 20–25 Kcal/kg/day in the acute flow phase and 25–30 Kcal/kg/day in the anabolic flow phase is preferred for measurement of calorie requirements. For overweight and obese patients, ideal body weight: 0.9 × height (cm) − 100 (male) (or − 106 (female) is suggested as a reference weight [26]. To avoid overfeeding, the EN target will be prescribed within 3 days in patients with high nutritional risk and within 7 days in patients with low nutritional risk according to the modified Nutrition Risk in Critically Ill (NUTRIC)² score. The flow charts in Figs. 2 and 3 will be used for initiation and continuation of EN.

In the intervention group, patients will receive Lactocare (ZistTakhmir) capsules 500 mg every 12 h for 14 days. Each capsule contains Lactobacillus casei 1.5 × 10⁹ colony-forming units (CFU), Lactobacillus acidophilus 1.5 × 10¹⁰ CFU, Lactobacillus rhamnosus 3.5 × 10⁹ CFU, Lactobacillus bulgaricus 2.5 × 10⁸ CFU, Bifidobacterium breve 1 × 10¹⁰ CFU, Bifidobacterium longum 5 × 10⁸ CFU, Streptococcus thermophilus 1.5 × 10⁸ CFU, and fructooligosaccharides (FOS). The probiotics capsule will be given through the NGT, separately from gavage, after feeding. Patients in the control group will receive a placebo capsule, which contains only sterile maize starch and is similar to probiotic capsules. The liquid preparations ready for gavage through the NGT are also similar in color and odor.

The pharmaceutical company will provide synbiotic and placebo capsules in distinct boxes identified as A or B. Synbiotic capsules can be stored at room temperature for 2–3 weeks but the best condition for keeping this product is in the refrigerator at 2–8 °C. Unused study products will be returned to the company supplying them.

Concomitant interventions will be:

These drugs are routinely administered in critical care practice, so we will record and consider them as conflicting factors.

As the researcher will administer the capsules to the patients through the NGT, adherence assessment is not required.

The schedule of evaluations is shown in Table 1.

As shown in Fig. 2, calorie achievement goals are set according to the patients’ modified NUTRIC score. In everyday visits, we will evaluate GI signs and symptoms (e.g. vomiting, diarrhea, abdominal distention) and gastric residual volume (GRV). If there is no sign or symptom of intolerance and GRV is less than 250 ml, EN will be increased by 10%. Otherwise, we will approach as shown in Fig. 3. Energy homeostasis (calorie intake-estimated calorie requirement) will be recorded each day. Mid-arm circumference, which is an available anthropometric measurement tool, will be evaluated twice a week. As all patients receiving EN should be monitored for some clinical and laboratory variables, we set our monitoring approach as reported by Berger MM, et al. [34], Monitoring nutrition in the ICU, Clinical Nutrition (2018). Concomitant medications, pressure ulcers, infectious complications, and other adverse events will be recorded every day. The APACHE II and Sequential Organ Failure Assessment (SOFA) will be scored on days 0, 8, and 15. Before and after the intervention, fasting blood and 24-h urine samples will be obtained. Urine urea nitrogen, 3-Methyl histidine, and creatinine will be measured in urine samples. Fasting blood glucose, insulin, C-reactive protein (CRP) and endotoxins will be measured in fasting blood samples.

Data will be analyzed using the intention-to-treat approach.

We did not find any similar study that has evaluated our primary objectives. So, we considered one of the main secondary objectives to estimate the required sample size. Mahmoodpoor and co-workers [35] reported the ICU stay in two study groups as 18.6 ± 8 and 11.6 ± 6.3 days. Considering alpha error of 0.05 and power of 80%, the required sample size allowing for 10% dropout is 20 patients in each group.

Data will be analyzed using SPSS for windows version 11.5 and MedCalc Statistical Software version 18.11.3 (MedCalc Software bvba, Ostend, Belgium; https://​www.​medcalc.​org; 2019). Descriptive (frequency, percentage, mean, standard deviation) and inferential analysis (student t test, paired sample t test, repeated measures analysis of variance (ANOVA)) will be performed. Any covariates will be controlled by ANCOVA or binary logistic regression. All tests will be two-tailed and a p value <0.05 will be considered as statistically significant.