Further, we investigated aberrant metabolites to find biomarkers for PD development and therapeutic targets based on the quantitative results. Unfortunately, most tryptophan metabolites in the urine samples did not present significant differences between control and PD patients, except for indole-3-acetic acid. Nonetheless, indole-3-acetic acid was reported for the first time for its aberrant urinary levels in PD patients and tentatively selected as a potential biomarker for PD. This study provides accurate quantitative results for 21 tryptophan metabolites in biological samples and will be helpful in revealing the pathological mechanisms of PD development, discovering biomarkers for PD, and further providing therapeutic targets for various PD symptoms. In the near future, to further investigate the relationship between gut microbial metabolites and PD, we will employ studies on microbial metabolites using plasma and stool samples from control and PD patients.Although Parkinson’s disease (PD) is a representative neurodegenerative disorder and shows characteristic motor impediments, the pathophysiological mechanisms and treatment targets for PD have not yet been clearly identified. Since several tryptophan metabolites produced by gut microbiota could pass the blood–brain barrier and, furthermore, might influence the central nervous system, tryptophan metabolites within the indole, kynurenine, and serotonin metabolic pathways might be the most potent targets for PD development. Furthermore, most metabolites are circulated via the blood, play roles in and/or are metabolized via the host organs, and finally are excreted into the urine. Therefore, profiling the overall tryptophan metabolic pathways in urine samples of patients with PD is important to understanding the pathological mechanisms, finding biomarkers, and discovering therapeutic targets for PD. However, the development of profiling analysis based on tryptophan metabolism pathways in human urine samples is still challenging due to the wide physiological ranges, the varied signal response, and the structural diversity of tryptophan metabolites in complicated urine matrices. In this study, an LC–MS/MS method was developed to profile 21 tryptophan metabolites within the indole, kynurenine, and serotonin metabolic pathways in human urine samples using ion-pairing chromatography and multiple reaction monitoring determination. The developed method was successfully applied to urine samples of PD patients ( Parkinson’s disease (PD) is the second most prevalent neurodegenerative disorder and has characteristic motor symptoms, including tremors, muscle rigidity, and impaired mobility ,2. AlthoFurthermore, since PD patients generally suffer from gastrointestinal (GI) problems such as constipation, dysbiosis of the gut microbiota is suspected to enable various PD symptoms. The interaction between the gut microbiota and the host, called the gut–brain axis, has been increasingly considered in many studies for various diseases besides PD ,9,10,11.Since most gut microbial and host metabolites are absorbed and distributed via blood circulation, play a role in and/or are metabolized by the organs, and are finally excreted into urine , the devn = 20) and PD patients (n = 41). On the basis of quantitative results on tryptophan metabolites in the urine samples, altered metabolism and dysbiosis were analyzed to unveil PD’s symptoms and mechanisms. In this study, the comprehensive profiling results of the 21 tryptophan metabolites in urine samples are provided, and a potential biomarker for PD is proposed. This study will be helpful in unveiling the pathophysiological mechanisms of PD and in understanding the motor and non-motor symptoms of PD.Therefore, to investigate the crosstalk between the gut microbiota and host in PD patients, we developed an LC–MS/MS method to simultaneously determine 21 tryptophan metabolites, particularly focused on indole metabolites, in human urine. The developed method was applied to 61 urine samples from a control group  labeled with deuterium were used to compensate for distorted detection results caused by the loss of analytes during sample preparation, variability in instrumental detection, or other unexpected errors. These ISs were selected on the basis of their structural characteristics and commercial availability. The overall validation parameters are summarized in The linearity of the analytes was investigated using calibration curves, which were established by analyzing artificial urine samples spiked with eight calibration points in six replicates. All determination coefficients were higher than 0.99, as shown in The stability results based on different storage and working conditions provided important information for working conditions. Overall, the analytes were relatively stable under refrigerator or autosampler conditions for a period of 3 days. Therefore, all analyte chemicals and samples were stored in a refrigerator or autosampler until instrumental analysis. Furthermore, although all analytes were stable during three freeze–thaw cycles, all samples were analyzed within 1 day after the thawing process. The matrix effect results were found to be in the range of 85.5–104.3%, showing that the analytes were not influenced by any significant matrix interference . Overalln = 41) and controls (n = 20). As shown in p-values of the 21 tryptophan metabolites in human urine from PD patients and controls are summarized in p-values of differences in urinary target metabolite levels between PD patients and controls were obtained via the Mann–Whitney U test using R 4.1.3 .Using the established method, the levels of 21 tryptophan metabolites were investigated in human urine from PD patients  was selected as a potential biomarker for PD in human urine . All authentic standards and chemicals were of analytical grade or better. All target metabolites  were purchased from Sigma-Aldrich . Isotopically labeled internal standards (ISs), kynurenic acid-dAll organic solvents were extra pure and of analytical grade for column chromatography. Analytical-grade acetonitrile (ACN) and methanol (MeOH) were obtained from Honeywell . Ethanol (EtOH) was purchased from J.T. Baker . Hydrochloric acid (HCl), sodium hydroxide (NaOH), dimethylsulfoxide (DMSO), and heptafluorobutyric acid (HFBA) (purity  ≥  98%) were purchased from Sigma-Aldrich . The acetonitrile and methanol used in the LC analysis were filtered through a 0.45 µm membrane filter and degassed for 10 min. De-ionized water (D.W.) was supplied using a Millipore Direct-Q3 purification system from the Millipore Corporation , then filtered through a 0.2 µm membrane filter and degassed for 10 min prior to use.v/v) and MeOH .Stock solutions of each deuterated internal standard and unlabeled analyte standard were prepared in different solvents due to the varying analyte solubilities at a concentration of 1 mg/mL. Most standard stock solutions were prepared in methanol. Tryptamine, tryptophol, and indole-3-propionic acid were dissolved in ethanol; kynurenic acid, kynurenine, and 3-hydroxy kynurenine were dissolved in 1 M HCl; xanthurenic acid was dissolved in 0.5 M NaOH; and indole-3-acetamide, indole-3-lactic acid, and indole acrylic acid were prepared in DMSO. These stock solutions were stored at −20 °C until use and were stable over a period of six weeks. Standard working mixtures were diluted with methanol, dried under a gentle stream of nitrogen gas, and re-dissolved in HFBA in de-ionized water  and patients with Parkinson’s disease (n = 41) were provided by Kyung Hee University Hospital, Seoul, Republic of Korea. Urine samples were collected via routine medical examination procedure (after fasting for at least 8 h). Detailed sample information and the ages and genders of the participants are listed in All human urine samples from the control group . The sample mixture was vortex-mixed for 30 s and centrifuged at 12,000 rpm for 10 min. After centrifugation, the supernatant (50 μL) was transferred into a 1.5 mL centrifuge tube. For deproteinization, 50 μL of MeOH was added to the sample; the mixture was vortex-mixed for 30 s and centrifuged at 12,000 rpm for 5 min. After centrifugation, the supernatant (80 μL) was dried under a gentle stream of nitrogen gas. The dried residue was re-dissolved in 40 μL of a mixture of HFBA in de-ionized water  and MeOH , followed by injection (10 μL) into the UPLC–MS/MS system. The overall analytical flow used to profile tryptophan metabolites in human urine samples is depicted in Collected urine samples were treated according to the method in a previous study  with minThe UPLC–MS/MS system used was a Waters UPLC H-class system  coupled to an API 3200 triple quadrupole mass spectrometer . v/v) and B . The column temperature was set to 40 °C. Gradient elution was performed as follows: 5% mobile phase B for 0.0–4.0 min, 5–25% B for 4.0–6.0 min, 25% B for 6.0–11.5 min, 25–80% B for 11.5–17.0 min, 80–100% B for 17.0–18.0 min, 100% B for 18.0–22.0 min, 100–5% B for 22.0–22.1 min, and 5% B for 22.1–25.0 min for re-equilibration. The flow rate and injection volume were set to 200 μL/min and 10 μL, respectively. Chromatographic separation was accomplished using a Waters ACQUITY UPLC BEH C18  reversed-phase column with mobile phase A . Samples were analyzed using Turbo V electrospray ionization (ESI) in positive ion multiple reaction monitoring (MRM) mode. All MS parameters were optimized by direct syringe infusion. The mass spectrometric conditions were set as follows: ion spray voltage (IS), 5200 V; curtain gas (CUR), 20 psi; nebulizer gas (GS1), 50 psi; auxiliary gas (GS2), 50 psi; temperature (TEM), 400 °C; and collision-activated dissociation gas (CAD), 5. Nitrogen was used as the nebulizing and collision gases. Quantitative and qualitative MRM transition ions were selected as the most and second-most abundant product ions, respectively. The optimized MRM parameters are summarized in 2). The precision and accuracy were evaluated using intra- and inter-day tests, which were validated using QC samples at high, medium, and low concentrations in six replicates. The recovery was calculated as / × 100 (%)). The matrix effect (ME) was calculated as /(area of standard solution) × 100 (%)). The stability of each compound was assessed using spiked samples at high, medium, and low concentrations under different storage conditions (room temperature or refrigerated at 4 °C) and working conditions (3 freeze–thaw cycles) before processing.The developed analytical method was validated in accordance with the bioanalytical validation guidelines provided by the US FDA . The metn = 20) and PD patients (n = 41). Since the individual metabolite levels in urine samples vary markedly according to individual status, it is difficult to identify potential urinary biomarkers for PD. However, according to the quantitative results for the 21 tryptophan metabolites in the urine samples, the urinary IAA level was significantly different between the two groups (p < 0.05). Although the causes of this increased urinary IAA level could not be identified, we cautiously suggest that indole metabolism pathways and the gut microbiota related to indole metabolism are distorted in PD patients. Furthermore, we speculate that an increased urinary IAA level might be a biomarker for PD, assuming that increased IAA in PD can be reconfirmed in subsequent studies on other biological samples and large-scale cohorts. In conclusion, this study provided a useful LC–MS/MS method to determine 21 tryptophan metabolites in biological samples in order to discover potential biomarkers and could be helpful in unveiling the pathophysiological mechanisms of PD.In this study, the levels of 21 tryptophan metabolites were determined in human urine using LC–MS/MS to investigate the unknown mechanisms of PD. Using HFBA, all target analytes were detected in positive ESI mode. Based on the quantitative and qualitative fragment ions, MRM transitions were established to sensitively and selectively determine the levels of the 21 tryptophan metabolites in human urine. The developed method was successfully applied to 61 human urine samples from a control group (


Bifidobacterium infantis has special abilities to utilise human milk oligosaccharides. Hence we hypothesised that probiotic supplements containing B. infantis may confer greater benefits to preterm infants than probiotic supplements without B. infantis.B. infantis and those that did not contain B. infantis.A systematic review with meta-analysis was conducted according to standard guidelines. We selected RCTs evaluating probiotics compared to placebo or no treatment in preterm and/or low birth weight infants. Probiotic effects on Necrotizing Enterocolitis (NEC), Late Onset Sepsis (LOS) and Mortality were analysed separately for RCTs in which the supplemented probiotic product contained n = 14,606), of which 16 used probiotics containing B. infantis (Subgroup A) and 51 RCTs did not (Subgroup B) Meta-analysis of all RCTs indicated that probiotics reduced the risk of NEC, LOS, and mortality. The subgroup meta-analysis demonstrated greater reduction in the incidence of NEC in subgroup A than subgroup B .67 RCTs were included (B. infantis may be more beneficial for preterm infants. Well-designed RCTs are necessary to confirm these findings.These results provide indirect evidence that probiotic supplements that include Evidence is emerging that beneficial effects of probiotics are species and strain specific.B. infantis supplementation provides an advantage to preterm infants.This systematic review analyses if B. infantis in preterm infants.This is the first systematic review evaluating the effects of probiotics containing B. infantis may be more beneficial for preterm infants. These results will help in guiding future research and clinical practice for using B. infantis as a probiotic in preterm infants.The results of this systematic review provides indirect evidence that probiotics that include  An important risk factor for NEC and sepsis in preterm infants is gut dysbiosis.2 Systematic reviews of randomised controlled trials (RCTs) and non-randomised studies have shown that probiotic supplementation reduces the risk of NEC (≥Stage II), LOS, and mortality in preterm infants.3 The benefits of probiotics relate to their ability to improve the gut barrier, modulate the immune system and attenuate gut dysbiosis.1 Probiotics have been shown to reduce the relative abundance of pathogens in the gut through various pathways, including blocking the receptors and competing for nutrients.4Probiotics are live microorganisms that when administered in adequate amounts, could confer beneficial effects on the host.5 data on individual probiotic species and strains is important for guiding clinical practice and research.Considering that probiotic effects are considered species and strain specific,6 Among the bifidobacteria, Bifidobacterium longum subspecies infantis (B. infantis) is considered as an important gut symbiont, especially in infancy. It is considered as a champion coloniser of the gut due to its properties for the consumption of human milk oligosaccharides (HMOs).7 It may have a competitive advantage against other bacteria, allowing increased colonisation and resulting in fewer luminal pathogens.8B. infantis promotes maturation of the innate immune response9 and improves the anti-inflammatory properties through the production of tryptophan metabolite, indole-3-lactic acid (ILA).10 Given these properties, we hypothesised that supplementation with probiotics containing B. infantis will be more beneficial in preterm infants than those without this sub-species of bacteria.During the early human development, bacteria belonging to the genus Bifidobacterium play an important role.3 has addressed this specific question. There are no RCTs in preterm infants that have compared B. infantis versus placebo. In addition, apart from the small RCT by our group,11 there are no RCTs that have compared supplementation with probiotics containing B. infantis versus probiotics not containing B. infantis.To our knowledge, no systematic review, including the latest systematic review and the network meta-analysisB. infantis versus placebo/no probiotics; Subgroup B: Probiotics not containing B. infantis versus placebo/no probiotics.Hence, we conducted a systematic review that had two subgroups: Subgroup A: Probiotics containing 12 and the PRISMA  statement13 were followed for undertaking and reporting this systematic review and meta-analysis.Guidelines from the Cochrane Neonatal Review Group,We selected RCTs evaluating probiotics for the prevention of morbidity or mortality in preterm  and/or low birth weight (birth weight <2500 g) infants. We excluded studies that enrolled term infants. Non-randomised studies, narrative reviews, systematic reviews, case reports, letters, editorials, and commentaries were excluded but read to identify potentially eligible studies.Preterm infants born before gestation <37 weeks, low birth weight (<2500 g), or both.We included studies assessing enteral administration of any probiotic commenced within the first week of life and continued for at least one week compared to placebo or no treatment. We excluded studies that used prebiotics or synbiotics (i.e. combination of a prebiotic with probiotics).14; (2) LOS defined as isolation of a pathogen from blood, cerebrospinal fluid, or a normally sterile body space after 48 h of birth; (3) All-cause mortality.These included (1) NEC ≥Stage II (Modified Bell’s criteria)www.thecochranelibrary.com, through December 2022), PubMed , EMBASE (Excerpta Medica dataBASE) via Ovid , EMCARE via OVID  databases were searched. We searched https://clinicaltrials.gov and ANZCTR  for ongoing RCTs. Grey literature was searched using Mednar (www.mednar.com). The reference lists of identified studies and key review articles were searched to identify additional RCTs. No language restriction was applied.The Cochrane Central Register of Controlled clinical trials (PubMed was searched using the following keywords:((((((((Probiotic) OR (Probiotics)) OR (Bifidobacteria)) OR (Bifidobacterium)) OR (Lactobacilli)) OR (Lactobacillus)) OR (Saccharomyces)) AND ((((((Preterm infant) OR (Preterm infants)) OR (premature infants)) OR (low birth weight infants)) OR (very low birth weight infants)) OR (extremely low birth weight infants))) AND . PubMed was also searched using relevant MeSH words. Other databases were searched using similar terminologies.Abstracts of the citations obtained from the initial broad search were read independently by two reviewers to identify potentially eligible studies. Full-text articles of these studies were obtained and assessed independently for eligibility by two reviewers, using the predefined eligibility criteria. Differences in opinion were resolved by a group discussion to reach a consensus. Multiple publications of the same study were excluded to avoid duplication of the data.Two reviewers independently extracted the data using a standardised data collection form. Discrepancies were resolved by discussion and consensus among all authors.12 Two reviewers independently assessed the ROB in all domains including random number generation, allocation concealment, blinding of intervention and outcome assessors, completeness of follow up and selectivity of reporting. For each domain, the ROB was assessed as low, high or unclear.ROB was assessed using the Cochrane “Risk of Bias Assessment Tool”.Meta-analysis was performed using statistical software, STATA (Version 17.0). Since heterogeneity was expected we used random-effects model (REM) model for meta-analysis. Fixed effect model (FEM) was also used to assure that the choice of the model did not influence the results. Since all outcomes of interest were binary, we used relative risk (RR) and 95% CI to summarise their results.I2 statistic and interpreted as per Cochrane handbook11 as follows: 0–40%: might not be important; 30–60%: may represent moderate heterogeneity; 50–90%: may represent substantial heterogeneity; 75–100%: considerable heterogeneity.Clinical heterogeneity was assessed and reported by summarising characteristics such as the study population, dose, and duration of probiotic supplementation. Statistical heterogeneity was estimated using the 15 Harbord’s,16 Begg’s17 and trim & fill plots.18To assess for any publication bias, we used Egger’s,19The key information about the quality of evidence, the magnitude of the effect of the intervention and the sum of available data on the main outcome was presented in the SOF table according to the Grades of Recommendation, Assessment, Development and Evaluation (GRADE) guidelines.n = 4), RCTs of prebiotics or synbiotics (n = 6), being conducted in full-term infants (n = 1), and not reporting our outcomes of interest (n = 6). Finally, 67 RCTs were included.86Initial broad search identified 2315 records, of which full texts of 84 potentially eligible studies were read in detail. Seventeen of these studies were excluded being non-RCTs , 16 (n = 4962) had used B. infantis as a component of the probiotic supplement.35 The remaining 51 (n = 9644) did not use B. infantis.86 The mean gestation and birth weight ranged from 25.4 weeks to 33.5 weeks and from 727 g to 2262 g, respectively. The duration of probiotic supplementation varied from a minimum of 2 weeks to until discharge/40 weeks corrected gestational age. The probiotic dose ranged from 18 × 106 (18 million) colony forming units (CFU) to 12 × 109 (12 billion) CFU/day. NEC, LOS and all-cause mortality were included as the outcomes in 63, 57 and 54 studies, respectively. The characteristics of the included studies are given in Table Of the 67 RCTs  studies were considered to have “low ROB” on the domain of random sequence generation, 5 (31.2%) had “high ROB” for performance & detection bias. Fifteen (93.7%) studies and 12 (75.0%) showed “low ROB” for attrition and reporting bias, respectively. Among the 51 studies that used probiotics without B. infantis, 36 (70.5%) and 28 (54.9%) were considered “low ROB” in the selection bias category. 24 (47%) studies had “unclear risk” or “high risk” ROB for blinding. For the attrition and reporting bias categories, 44 (86.2%) and 35 (68.6%) studies showed “low ROB”. Details of the ROB analysis are given in Table Of the 16 studies that used probiotics containing B. infantis versus those that used probiotics without B. infantis.The effects of the intervention were compared between the studies that used probiotics containing I2 = 0% ; RR 0.59 (CI 0.50–0.70); n = 3626) that used probiotics containing B. infantis (Subgroup A) showed significant reduction in the incidence of NEC  that used probiotics without B. infantis (subgroup B) also found significant reduction in the incidence of NEC  and Begg’s (p = 0.577) tests showed no publication bias exists but the Egger’s test (p = 0.010) did. Further, trim & fill analysis imputed 9 potentially missing studies ; RR 0.84 (CI 0.76–0.94); %); RR 0.n = 4123) that used probiotics containing B. infantis (subgroup A) showed that probiotics reduced the incidence of LOS  that used probiotics without B. infantis (subgroup B) also found reduction of LOS  and Begg’s (p = 0.188) tests showed no publication bias exists but the Egger’s test (p = 0.088) did. Further, trim & fill analysis imputed 5 potentially missing studies ; RR 0.78 (CI 0.67–0.91); n = 4292) that used probiotics containing B. infantis (subgroup A) showed significant reduction of all-cause mortality  that used probiotics without B. infantis (subgroup B) also found reduction of all-cause mortality , Begg’s (p = 0.560) and Egger’s tests (p = 0.100) showed that there is no publication bias. However, the trim & fill analysis imputed 5 potentially missing studies. Meta-analysis after incorporating the results of imputed studies found results that were similar to the primary analysis  ] Fig. .Fig. 5TrB. infantis was a component of the probiotic supplement, the overall GRADE of evidence was high for the outcomes of mortality and NEC and moderate for LOS  found that probiotic supplementation significantly reduced the risk of NEC≥ Stage II, LOS and all-cause mortality in preterm infants. Specific to our aim, the sub-group meta-analysis of RCTs that used probiotics containing B. infantis showed even more favourable results, especially for the prevention of NEC (≥Stage II). These results provide indirect evidence that probiotics that include B. infantis may be more beneficial in preterm infants than those not including B. infantis.Our systematic review that included 67 RCTs (B. infantis was associated with a significant reduction in NEC (≥Stage II) and NEC-related mortality.87 The B. infantis cohort had a 73% reduction of NEC (≥ stage II) compared with the no B. infantis cohort .Our results are supported by a recent non-randomised study by Tobias et al, involving 483 VLBW infants. Supplementation of B. infantis administration on gut microbiota, nosocomial acquired antibiotic resistance and enteric inflammation in preterm infants with gestation <32 weeks and/or birth weight <1500 g.88 Infants supplemented with B. infantis had lower enteric inflammation after adjusting for other clinical variables in multivariate modelling. In contrast, Kochjancic et al. reported that probiotic (B. infantis and Lactobacillus acidophilus) supplementation did not reduce the risk of NEC in neonates with duct-dependent congenital heart disease (CHD).89 The lack of benefits of probiotics may relate to the small sample size (n = 15) and retrospective design of the study, lack of concurrence of NEC and duct-dependent CHD, and the fact that the majority of infants were born at term.A prospective study by Nguyen et al. has evaluated the effect of B. infantis is important as probiotic effects are species and strain- specific. Ward et al. and LoCascio et al. reported that given their complex structure, the HMOs cannot be metabolised by the infant or most of the bacteria in the infant’s gut, as they lack the necessary enzymes for the purpose.91B. infantis has been shown to grow in vitro using HMO as the sole carbon source, reaching a cell density 3-fold higher than B. longum subsp. longum, B. breve, B. bifidum, and B. adolescentis. Low pH is a critical factor in preventing the invasion and overgrowth of harmful bacteria in the infant gut, a process known as colonisation resistance. Henrick et al. reported that B. infantis supplementation significantly lowered faecal pH in breastfed infants compared to controls.92 Underwood et al. reported that HMO metabolism by B. infantis produces short-chain fatty acids (SCFA), such as acetate, which play an important role in nutrition and intestinal and immune development, facilitate direct binding to intestinal cells, and stimulate anti-inflammatory/inhibits pro-inflammatory cytokine release by intestinal cells.93 Meng et al. reported that B. infantis contributes to maintaining of gut barrier integrity through indole 3-lactic acid (ILA), a metabolite of tryptophan, and may protect gut epithelium by activating the aryl hydrogen receptor, which can further promote intestinal immune function.10 In addition to their role in the gut, SCFAs produced by B. infantis can enter circulation and directly affect the adipose tissue, lungs, brain, and liver, inducing overall beneficial metabolic effects.94 Animal studies by Bergmann et al. suggested that B. infantis can potentially protect against excessive intestinal inflammation which is implicated in the pathogenesis of NEC in preterm infants.95 Given that probiotics are live organisms, a major concern is the risk of sepsis due to the administered probiotic organism. Although there are few case reports of bacteraemia caused by the Bifidobacteria,97 it is reassuring to note that none of the RCTs included in our review that used probiotics reported probiotic related sepsis. However, current evidence is limited for estimating the risk of probiotic sepsis.99 In 2007, the European Food Safety Authority (EFSA) assigned qualified presumption of safety (QPS) status to the bacterial species B. longum, which includes subspecies infantis, indicating that this taxonomic group does not carry safety concerns.100 The QPS status, which applies to all strains of B. infantis indicates that none of these has been associated with human clinical disease. However, it should not lead to complacency, and constant microbiological surveillance is essential to identify and treat sepsis that may occur due to the administered probiotic organism.Discussing the physiological characteristics of B. infantis in preterm infants. Our results will help in guiding research for using B. infantis as a probiotic in preterm infants. This is also one of the largest systematic reviews of probiotics in preterm infants involving 14,606 preterm infants, which is close to the recent network meta-analysis (3) and 3800 infants more than the Cochrane review.101To our knowledge, ours is the first systematic review related to B. infantis and the remaining 51 RCTs used a probiotic that did not contain B. infantis, it resulted in a large discrepancy between the number of participants between these two groups. Furthermore, none of the included RCTs used B. infantis as the sole probiotic. Instead, they used a mixture of probiotic organisms with variable doses of B. infantis. Thus, the observed benefits cannot be attributed definitively to B. infantis.Our systematic review has some limitations. Since only 16 RCTs used probiotics containing B. infantis is a component of the probiotic product compared to studies in which B. infantis is not a component. However, given the limitations to the evidence, adequately powered RCTs are necessary to confirm the benefits and safety of B. infantis in preterm infants. Such RCTs could compare (1) B. infantis versus Placebo or (2) B. infantis as a component of a multi-strain probiotic product versus the same multi-strain probiotic but without B. infantis.In conclusion, our systematic review of RCTs provides indirect evidence that the beneficial effects for the prevention of NEC are more pronounced if PRISMA_checklist


Leishmania biology field in recent decades, the mechanisms involved in DC/NK-mediated control of Leishmania sp. pathogenesis as well as the cellular and molecular players involved in such interaction remain unclear. The present study sought to investigate canonical pathways associated with CL arising from Leishmania braziliensis infection. Initially, two publicly available microarray datasets of skin biopsies from active CL lesions were analyzed, and five pathways were identified using differentially expressed genes. The “Crosstalk between DCs and NK cells” pathway was notable due to a high number of modulated genes. The molecules significantly involved in this pathway were identified, and our findings were validated in newly obtained CL biopsies. We found increased expression of TLR4, TNFRSF1B, IL-15, IL-6, CD40, CCR7, TNF and IFNG, confirming the analysis of publicly available datasets. These findings reveal the “crosstalk between DCs and NK cells” as a potential pathway to be further explored in the pathogenesis of CL, especially the expression of CCR7, which is correlated with lesion development.Skin ulcers of cutaneous leishmaniasis (CL) are characterized by a localized inflammatory response mediated by innate and adaptive immune cells, including dendritic cells (DC) and natural killer (NK) cells. Bidirectional interactions between DCs and NK cells contribute to tailor leishmaniasis outcome. Despite advances in the  Leishmania whose main vectors are phlebotomine sandflies. This disease is considered a worldwide public health problem, as it is present in 102 countries, placing approximately 1 billion people at risk of infection, with an estimated 12 million people currently infected. Each year, approximately two million new cases are registered, resulting in 70,000 deaths [Leishmaniasis is a chronic inflammatory disease caused by protozoan parasites of the genus 0 deaths ,4,5.Leishmania (Viannia) braziliensis [L. braziliensis infection, the Th1 response is fundamental for parasite elimination and lesion resolution; however, this same immune response promotes tissue damage [Cutaneous leishmaniasis (CL) is the most common form of human leishmaniasis, accounting for approximately 95% of cases, on average . In Braziliensis  is the miliensis . It has e damage ,9,10,11.e damage ,13. Bothe damage ,11,12,13DCs are potent professional antigen-presenting cells (APCs) that effectively link innate and adaptive responses. Furthermore, DCs are a heterogeneous cell population and classified according to origin, phenotype, tissue distribution and function . Under rNK cells constitute a subset of granular lymphocytes that play a dual role in immunity. These cells can orchestrate cytotoxic or immunomodulatory processes mainly involved in the recognition and elimination of stressed cells from different types: infection, tumor-related transformations, and physical damage ,16,17. TAbundant evidence suggests that NK cells regulate the functions of other cells via the production of cytokines and chemokines and/or direct cell–cell contact (receptor–ligand interactions) ,24. ThisLeishmania braziliensis. The investigation of this pathway in the context of parasite infections has been previously unclear, and the study aims to shed light on this aspect, providing new insights and potential strategies for the management of this infection. By examining the transcriptome profiles, the study aims to uncover specific molecular pathways and mechanisms involved in CL, which could be targeted for the development of novel therapeutic interventions. The findings from this research have the potential to broaden our understanding of the host-parasite interaction in CL and pave the way for innovative approaches to tackle this challenging infectious disease.The present study sought to analyze transcriptome datasets to identify canonical pathways associated with the innate immune response in cutaneous leishmaniasis (CL) lesions caused by www.ncbi.nlm.nih.gov/geo/ (accessed on 15 February 2022) using the following search terms: ‘human’, ‘biopsy’, ‘skin’ and ‘Leishmania braziliensis’. The dataset published by Novais et al. (GSE55664) [Transcriptomic data sets were obtained from the Gene Expression Omnibus (GEO) database (SE55664)  was geneSE55664)  was genemev.tm4.org/ (accessed on 15 March 2022)) microarray data analysis tool to generate heat maps representing significantly DEGs according to the following criteria: a previously evaluated FDR adjusted p-value < 0.05, and fold change of ≤−2 or ≥+2. The default parameters of MEV were used. The hierarchical clustering considered was the ‘Gene tree’ and ‘Sample tree’ and the current metric by ‘Euclidean distance’. The normalization was performed by ‘gene’ and ‘sample’ and the set color scale limits according to the displayed default value.Raw gene expression data were analyzed using the Multi Experiment Viewer (www.qiagenbioinformatics.com (accessed on 15 March 2022) to identify pathways related to CL. The search term “Immune response” was applied to identify specific pathways of interest, considering only DEGs in each dataset.Each dataset (GSE55664 and GSE63931) was analyzed by Ingenuity Pathway Analysis (IPA) . After, 140 μL of chloroform  was added and vigorously vortexed followed by incubation for 5 min at room temperature. The aqueous phase was recovered, mixed with 500 μL 70% ethanol, and RNA was obtained using the RNeasy mini kit  in accordance with the manufacturer’s instructions. RNA yield was assessed using a NanoDrop1000 spectrophotometer .To validate the results of our DEG analysis, eight genes were selected for relative quantification considering their fundamental importance to the pathway “Crosstalk between DC and NK cells” identified by IPA: Surface receptors TLR4, CD40 and CCR7; Cytokines produced by DCs TNF, IL15, IL6; TNFRSF1B, responsible for recognizing TNF; and IFNG, the cytokine produced by NK cells.−ΔΔCt method, and results were normalized using the ACTB housekeeping gene.A total of 1 μg of RNA was converted into complementary DNA (cDNA) using a SuperScript™ III Reverse Transcriptase kit (Invitrogen Cat: 18080085). Quantitative real-time PCR was performed using ABI7500 Real-Time PCR system  with 10 uL of SYBRGreen Real-Time PCR Master Mix. All relevant primers were synthesized by Integrated DNA Technologies. The relative expression of these genes was evaluated using the 2This study was conducted in accordance with the principles of the Declaration of Helsinki and local ethical guidelines. This study was approved by the Ethics Committee of Instituto Gonçalo Moniz . All patients who participated in the research provided written consent for sample collection (TCLE—Free and Informed Consent Form) and subsequent analysis.p-values < 0.05 were considered significant. For comparisons between groups, the Shapiro–Wilk test was used to detect data normality. For the evaluation of 2 groups, both with a normal distribution, Student’s t-test was applied, and for samples with non-normal distribution, the Mann–Whitney test was used. In analyzes with more than 2 groups, we used the analysis of variance test (ANOVA) for samples with normal distribution and Kruskal–Wallis followed by Dunn’s post-test for samples with non-normal distribution. Graphs were presented with a 95% confidence interval for the mean  or median with interquartile deviation . The results were represented with the median values obtained in each condition. Correlation analyses were performed by Spearman test and the matrix summarizes the statistically significant correlations. The correlation matrix is reordered according to the correlation coefficient using “hclust” method (http://www.sthda.com/english/rsthda/correlation-matrix.php (accessed on 20 May 2022). The 2−ΔΔCt method is a widely used approach in quantitative real-time PCR (qPCR) data analysis. It is commonly used to determine the relative gene expression levels between different samples or experimental conditions.GraphPad Prism 8  was used for statis-tical evaluation and p < 0.05 and ≤−2 FC ≥ +2) in the GSE55664 dataset: 1243 were up-regulated (yellow) and 1125 were down-regulated (blue). In the GSE63931 dataset, 6152 DEGs were identified: 2851 up-regulated (yellow) and 3301 down-regulated (blue)  A. The red (blue) B and illp values, a high degree of similarity was observed between pathways related to immune response in each dataset: Communication between Innate and Adaptive Immune Response; Crosstalk between Dendritic Cells and Natural Killer Cells; Granulocyte Adhesion and Diapedesis and Interferon Signaling  in the GSE55664  were obtained from a different endemic area, as well as healthy controls (n = 7) from a non-endemic area. The relative expression of TLR4  infection. Comparisons of gene expression among different tissues in WNV infection in mice identified the ‘Crosstalk between Dendritic Cells and Natural Killer Cells’ as important in this infection, suggesting a new antiviral therapy against flavivirus infections . AnotherLeishmania infection. Studies have shown that L. major, L. infantum and L. amazonensis exhibit a mechanism of immune evasion through the inhibition of DC activation, migration and immune function, which interferes in the induction of adaptive immune responses [Although the role of crosstalk between DCs and NK cells in leishmaniasis remains poorly understood, many studies have demonstrated the importance of DCs and NK cells in esponses ,43,44,45esponses ,47.L. amazonensis-infected DCs with resting NK cells results in increased activation of both cells, beyond augmented production of proinflammatory cytokines, including IFN-γ. In this same study, murine DCs infected by L. braziliensis amastigotes became activated, which was not the case in L. amazonensis infection [L. major infection via crosstalk with NK cells, leading to their activation and release of IFN-γ [It is known that interactions between DCs and NK cells can result in the amplification of inflammatory activity by both cells . In vitrnfection . Moreoveof IFN-γ . Our res


Several studies have evaluated the effects of tributyrin on sow reproductive performance; however, none of these studies have investigated the effects of tributyrin on sow gut microbiota and its potential interactions with immune systems and milk composition. Therefore, we speculated that tributyrin, the combination of butyrate and mono-butyrin without odor, would reach the hindgut and affect the intestinal microbiota composition and play a better role in regulating sow reproductive performance, gut flora, and health.Thirty sows (Landrace × Yorkshire) were randomly divided into two groups: the control group (CON) and the tributyrin group (TB), which received basal diet supplemented with 0.05% tributyrin. The experimental period lasted for 35 days from late pregnancy to lactation.Lactobacillus increased in sows and piglets, but the relative abundance of Subdoligranulum and Eubacterium_fissicatena_group decreased in piglets in the TB group.The results showed that TB supplementation significantly shortened the total parturition time and reduced the diarrhea rate in suckling piglets. On day 20 of lactation, the milk fat and protein levels increased by 9 and 4%, respectively. TB supplementation significantly improved the digestibility of dry material, gross energy, and crude fat in the sow diet, but had no significant effect on crude protein digestibility. Furthermore, TB supplementation increased the levels of IL-10, IL-6, and IgA in the blood of weaned piglets, but had no effect on maternal immunity. Analysis of the fecal microbial composition revealed that the addition of TB during late gestation and lactation increased the microbiota diversity in sows and piglets. At the phylum level, sows in the TB group had a slight increase in the relative abundance of Bacteroidota and Spirochaetota and a decrease in Firmicutes. At the order level, the relative abundance of Lactobacillales was increased in piglets and sows, and the TB group showed increased relative abundance of Enterobacterales and significantly decreased relative abundance of Oscillospirales in piglets. At family level, the relative abundance of Lactobacillaceae, Oscillospiraceae, and Christensenellaceae increased in sows, and the relative abundance of Enterobacteriaceae and Lactobacillaceae increased in piglets. At genus level, the relative abundance of In conclusion, tributyrin supplementation shortened the farrowing duration and reduced the diarrhea rate of piglets by improving the inflammatory response and composition of gut microbiota in piglets and sows. It is oin vivo to prevent fatal infection in weaned pigs, and improves the growth performance of piglets  and was conducted in accordance with the Guide for the Care and Use of Laboratory Animals of the National Research Council.n = 15 each): the control group (CON) was fed a basic diet and the tributyrin group (TB) was fed a basic diet supplemented with 500 g/t tributyrin . The diet was designed according to the experimental design and nutritional needs of the Thirty Landrace × Yorkshire sows with similar parity and backfat thickness (15.59 ± 2.63) were selected and randomly divided into two groups . The total litter size, number of piglets born alive, weight after birth, and weight on day 21 were recorded, and the weaning survival rate, daily weight gain, and litter weight gain were calculated. During the entire trial period, the fecal morphology of the sows was checked and recorded at a fixed time every day, and the degree of constipation was scored according to 2.3.g at 4°C for 10 min, and serum was separated and immediately stored at −20°C for later analysis. Colostrum samples were collected from the fourth or fifth pair (any nipple or more nipples) breast after the birth of the fifth piglet. On day 20 of lactation, milk samples were collected from sows by injecting 1.0 mg of oxytocin into the auricular vein and stored at −20°C for later analysis.Feed samples were collected by quartering and were hermetically stored at −20°C. On day 14 of lactation, fecal samples of sows were collected for five consecutive days and stored at −20°C for nutrient digestibility by endogenous indicator method. After collection, the fecal samples from each pig were mixed well, and approximately 200 g of the samples were collected by quartering and hermetically stored at −20°C. The fecal samples of sows and piglets were collected on day 21 of lactation for microbial analysis. Twelve litters were selected for each treatment, and a piglet close to the average weight were selected for each litter. The feces of sows and piglets in all treatment groups were collected by rectal massage method and stored hermetically at −20°C. On day 0 and 20 after parturition, blood samples (10 mL) were collected by ear venipuncture. On day 21 of lactation, blood from lactating piglets (5 mL) was collected from the anterior vena cava. All blood samples were centrifuged at 3,000× 2.4.CF), acid detergent fiber (ADF), and neutral detergent fiber (NDF) in the feed and fecal samples were measured according to Dry matter (DM), crude protein (CP), crude fat  were determined using HITACHI 3100 Automatic Analyzer . Plasma concentrations of reproduction-related regulatory hormones  in sows were determined using commercial ELISA kits , according to the manufacturer’s protocol.2.6.The concentrations of fat, protein, lactose, total solids, and urinary nitrogen in colostrum and milk were determined using an automatic milk composition analyzer . The concentrations of interleukin-A (IgA) and interleukin-A (IgM) were determined by the commercial ELISA kits .2.7.Interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-G (IgG), IgA, interleukin-M (IgM), Interleukin-2 (IL-2), and Interleukin-10 (IL-10) levels in the blood of sows and piglets were determined using commercial ELISA kits  and an Absorbance Microplate Reader (SpectraMax190).2.8.® Soil DNA Kit . Using this DNA as a template, the V3-V4 hypervariable regions of the bacterial 16S rRNA gene were amplified using primers F338 (5′-ACTCCTACGGGAGGCAGCAG-3′) and R806 (5′-GGACTACHVGGGTWTCTAAT-3′). An AxyPrep DNA Gel Extraction Kit was used to purify the amplicons. Purified amplicons were paired-end sequenced on an Illumina MiSeq PE300 platform/NovaSeq PE250 platform . Operational taxonomic units (OTUs) with a 97% similarity cut-off were clustered, and chimeric sequences were identified and removed. The taxonomy of the representative sequence of each OTU was analyzed  but showed no significant difference in litter size, live litter size, primary weight of piglets, or constipation score (p > 0.05). However, the constipation rate was reduced by 31% (p > 0.05).As shown in 3.2.p > 0.05).As shown in 3.3.p > 0.05), and significantly reduced rate of diarrhea in lactating piglets (p < 0.05).As shown in 3.4.p = 0.089), 4%, and 7.6%.As shown in 3.5.p = 0.092). In addition, blood triglycerides of piglets increased by 16% in the TB group.As shown in 3.6.p < 0.01), and crude fat (p < 0.05) in the sow diet, but had no significant effect on crude protein digestibility.As shown in 3.7.p = 0.058) and decreased INS in the blood on day 20 of lactation (p = 0.086).As shown in 3.8.p < 0.05) in the blood of 21-day-old lactating piglets with a tendency to increase the concentration of IL-10 (p < 0.10).As shown in 3.9.As shown in -Tissierellales (p < 0.01). As shown in p < 0.01) and Clostridiacea. As shown in Lactobacillus, significantly reduced the relative abundance of Romboutsia (p < 0.01). Therefore, the above results indicate that the gut microbiota composition of sows was profoundly altered during late pregnancy and lactation.Compared with the control, the microbiota of the fecal samples in the TB group showed a slight increase in the relative abundance of Bacteroidota and Spirochaetota, and a decrease in Firmicutes . As showp = 0.034). Similarly, the relative abundance of Enterobacteriaceae was also increased in the TB group (p = 0.032). As shown in Lactobacillus, Escherichia-Shigella, and Lachnoclostridium and decreased the relative abundance of Subdoligranulum and Eubacterium_fissicatena_group. Therefore, the above results indicate that the gut microbiota composition of piglets was profoundly altered during lactation.TB group , and thep = 0.020), and there had a tendency to increase in piglets (p = 0.094), compared to the CON group. As shown in Principal coordinate analysis (PCoA) revealed a significant increase in gut microbiota diversity following dietary tributyrin supplementation in sows  can be found in the article/The animal study was reviewed and approved by the Institutional Animal Care and Use Committee of Sichuan Agricultural University (SCAUAC202108-3).YL and ZM conceived and designed the study. YL, YW, ZM, LC, ZF, and SX performed the experiments and analyzed the data. BF, JL, LH, JZ, DW, and YZ performed the lab analysis. YL and DL wrote and edited the original draft. All authors have read and agreed to the published version of the manuscript.This study was funded by the National Key R&D Program of China (2021YFD1300202), the Sichuan Major Science and Technology Projects (2021ZDZX0009), and the Perstorp (Shanghai) Chemical Trading Co., Ltd. .YW was employed by the company Perstorp (Shanghai) Chemical Trading Co., Ltd.The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


At slaughter, samples of liver, muscle and blood serum were taken and underwent subsequent metabolomics profiling using a UHPLC-QTOF-MS system. A total of 6540 features could be detected. Twenty metabolites in the liver, five metabolites in muscle and thirty metabolites in blood serum were affected (p < 0.05) due to dietary treatments. In total, six metabolites could be reliably annotated and were thus subjected to subsequent univariate analysis. Reduction in dietary CP had minimal effect on metabolite abundance in target tissues of both RED and RED+RPMET bulls as compared to CON bulls. The addition of RPMET altered the hepatic anti-oxidant status in RED+RPMET bulls compared to both RED and CON bulls. Results exemplify nutrient partitioning in growing German Simmental bulls: bulls set maintenance as the prevailing metabolic priority (homeostasis) and nutrient trafficking as the second priority, which was directed toward special metabolic functions, such as anti-oxidant pathways.This study aimed to determine the metabolic response of growing German Simmental bulls fed rations low in crude protein (CP) supplemented with rumen-protected methionine (RPMET). In total, 69 bulls (on average 238 ± 11 days of age at start and 367 ± 25 kg of bodyweight) were assigned to three dietary treatments ( Beef supply chains have gained significant attention in the global discussions on livestock production. This public opinion is mainly derived from the fact that beef cattle have lower feed-to-food transformation efficiency as compared to both poultry and swine , meanwhiThis nutritional concept cannot be simply transferred to beef and dairy nutrition, since the fore stomach system synthesizes its own protein in the rumen . Thus, there is an incongruence between ingested and absorbed amino acid patterns, which impedes the determination of performance-limiting amino acids .Nevertheless, numerous studies imply that lysine and methionine might be the first-limiting amino acids in growing ruminants , more spMethionine is a sulfurized amino acid, which enters the one-carbon metabolism in its active form S-adenosylmethionine. As such, it donates its methyl group to an acceptor, thereby producing S-adenosylhomocysteine, which can be then hydrolyzed to homocysteine. This trans-methylation process, which is restricted to the liver, kidney, intestine and pancreas, is the first step of numerous subsequent metabolic pathways. It is followed by the trans-sulfuration pathway, which converts homocysteine into cysteine. Cysteine, in turn, serves as the intermediate metabolite for glutathione synthesis, an important antioxidant, or is oxidized into cystine, for instance. Other metabolic fates of cysteine are, e.g., the synthesis of taurine. Taurine, in turn, has several metabolic functions, such as antioxidant activity ,9,10.®, Adisseo, France). Reduction in dietary crude protein led to a decrease (p < 0.05) in growth performance over the whole experiment in both RED and RED+RPMET as compared to the CON group. The addition of RPMET did not relieve the RED+RPMET bulls from growth limitations, suggesting that under those feeding conditions, methionine was not firstly limiting for growth. However, the supplementation of RPMET led to an increase (p < 0.01) in serum methionine concentrations in RED+RPMET as compared to RED bulls (43.4 vs. 35.0 μmol/L) and hence, increased its metabolic availability [Protecting dietary methionine from ruminal degradation increases its metabolic availability in ruminants . This halability . Based oThe feeding experiment was conducted at the Bavarian State Research Center for Agriculture . All experimental procedures followed the guidelines of the German law under the German State and Directive 2010/63/EU of the Parliament and of the Council of 22 September 2010 on the protection of animals used for scientific purposes. Slaughter of the animals was conducted in accordance with the German law of animal protection of the German State and Council Regulation (EC) No. 1099/2009 of 24 September 2009 on the protection of animals at the time of killing.A feeding trial was conducted to harvest liver and muscle tissue and serum samples at the slaughter of the bulls. n = 23/group). Bulls were weighed at the beginning and immediately before slaughter as well as in regular intervals during the experimental period. The CON ration provided adequate metabolizable energy , crude protein (13.7%) and utilizable crude protein (15.7%), referring to the current national recommendations of German Simmental bull feeding [®, Adisseo SAS, Antony, France). The feeding trial lasted for 105 days on average. In brief, a total of 69 growing fattening German Simmental bulls  were allotted to a control group (CON) and two dietary treatment groups (Details on the feeding experiment were previously described .2 of the left lobe of the liver) and muscle (Musculus masseter) tissue samples of each bull were removed and directly frozen in a liquid nitrogen container to minimize metabolic degradation. Samples were stored in 1.5 mL cryotubes at −80 °C until assayed. Additionally, blood samples were collected in Vacuette tubes  during exsanguination. Vacuette tubes were inverted and centrifuged  and the serum samples were transferred to 1.5 mL cryotubes and then stored at −80 °C until further analysis.At the end of the feeding experiment, bulls were slaughtered over a time period of eight d and bulls from all three treatments were slaughtered on each date. At slaughter, liver  and the clear supernatants were transferred to 1.5 mL glass autosampler vials. Blood serum samples (200 μL) were diluted in the same extraction solvent used for tissue samples (1 mL). Muscle, liver and serum samples were thawed at 4 °C. Subsequently, 10 mg of each tissue (muscle and liver) sample were transferred into 2 mL FastDNA Lysing Matrix A tubes containing lysing matrix A for soft animal tissues . After adding 1 mL of an extraction solvent , samples were homogenized using FastPrep-24TM Classic  and incubated on ice (30 sec after each spin). Samples were then centrifuged ) and was performed with the following elution gradient: 100% B from 0 to 1.5 min, 60% B at 8 min and 20% B at 10 min to 11.5 min and 100% B at 12 to 15 min. A volume of 10 µL per sample was injected. The autosampler was cooled to 10 °C and the column oven was heated to 40 °C. A quality control (QC) sample was run after every ten samples . It consisted of a pool of all injected samples. MS settings in the positive mode were as follows: gas 1, 55; gas 2, 65; curtain gas, 35; temperature, 500 °C; ion spray voltage, 5500; declustering potential, 80. The mass range of the TOF MS and MS/MS scans were 50–2000 m/z. The collision energy was ramped from 15 to 55 V. MS settings in the negative mode were set as follows: gas 1, 55; gas 2, 65; cur, 35; temperature, 500 °C; ion spray voltage, −4500; declustering potential, −80. The mass range of the TOF MS and MS/MS scans was 50–2000 m/z. The collision energy was ramped from −15 to −55 V.Untargeted metabolomics analyses were carried out at the Bavarian Center for Biomolecular Mass Spectrometry . The untargeted analysis was performed using a Nexera UHPLC system (Shimadzu), which was coupled to a Q-TOF mass spectrometer  using the information-dependent acquisition (IDA) mode. A UPLC BEH Amide 2.1 × 100, 1.7 µm analytic column  with a 400 µL/min flow rate was used for separation of the biological samples. The mobile phase consisted of eluent A (5 mM ammonium acetate in water) and eluent B  and due to the addition of RPMET (RED vs. RED+RPMET). The SEM represents the standard error of the mean over the whole GLM. Statistical significance was declared at p ≤ 0.05.Univariate analysis was performed using SAS . Treatment group means of significantly regulated metabolites, which were obtained from XCMS analyses, were tested via normal distribution (Shapiro–Wilk’s test) and then analyzed using a general linear model (GLM) with treatment as fixed effect and a Student–Newman–Keuls (SNK) post-hoc test according to the SAS user’s guide . The p-vp < 0.05) dry matter intake  as compared to the standard group (9.43 kg/day in CON) over the whole duration of the experiment. This led to a reduction (p < 0.01) in digestible protein, metabolizable energy and nutrient intake in both RED and RED+RPMET groups as compared to the CON group. Consequently, growth rates were lower in RED and RED+RPMET bulls (p < 0.01) than in CON bulls. Considering the lower growth rates in both RED and RED+RPMET in a retrospective view, digestible protein supply in both RED and RED+RPMET met their requirements at 133 and 138%, respectively. Due to the specific addition of RPMET in RED+RPMET, these bulls had a higher (p < 0.01) intake of pre-cecal digestible methionine (26.2 g/day) as compared to RED bulls (18.8 g/day). Again, considering the lower growth rates in a retrospective view, methionine intake matched their daily requirements at 146% and 101%, respectively.Zootechnical results on the feeding experiment were previously described . In brieIn total, 930 and 527 features were identified in positive (P) and negative (N) ionization modes, respectively, in liver tissue. A total of 1122 and 747 features were identified in P and N ionization modes, respectively, in muscle tissue, and 1835 and 1379 features were identified in blood serum samples, respectively .p-value. Features depicted in ‘green’ show a match with MS2 spectra, whereas features in ‘violet’ lack a respective match. Each figure shows four volcano plots. In all figures  and five between RED and RED+RPMET (four detected in N mode and one detected in P mode), respectively .In muscle tissue, five metabolites were found to be statistically different between CON and RED+RPMET  .In total, 30 serum metabolites were significantly regulated, 29 between CON and RED+RPMET (13 detected in N mode and 16 detected in P mode) and one between RED and RED+RPMET (detected in N mode).p < 0.01) in hepatic carnosine (5.09 vs. 4.79), cystine (4.65 vs. 4.18) and taurocholic acid (4.69 vs. 4.31) and blood serum pyrrolidonecarboxylic acid (4.61 in CON vs. 3.97 in RED) concentrations. Additionally, the MS1 and MS2 spectra indicated the presence of L-leucine/L-isoleucine/norleucine with (p < 0.01) between CON and RED (4.37 vs. 4.13). Based on the retention time and the fragmentation pattern, we cannot differentiate between these amino acids and therefore combine them. The addition of RPMET (RED+RPMET) did not affect metabolite concentrations as compared to RED, with the exception of taurocholic acid  and cysteine glutathione disulfide .The transformation efficiency of human non-edible biomass into protein food derived from ruminants is relatively low compared to other species such as poultry and swine . HoweverHence, the objective of this study was to evaluate the effect of RPMET in reduced crude protein diets on metabolic pathways in growing German Simmental bulls under practical feeding conditions. As to our knowledge, this is the first study evaluating the metabolic response of German Simmental beef cattle that were fed diets reduced in crude protein supplemented with RPMET. Until 2023, there were 153 publications on metabolomics approaches in research with bovine species available . RecentlMetabolomics is an ‘omics‘ tool to quantify a global set of metabolites within biological samples. Hence, this analysis tool delivers the entirety of metabolic downstream products derived from genomic, transcriptomic and proteomic processes . These dp < 0.05) growth performance than CON bulls. Metabolomics analyses on a cellular level in muscle, blood and liver tissue revealed that there was hardly any difference in metabolite abundance between CON and both RED and RED+RPMET bulls. This exemplifies the hierarchical concept of nutrient partitioning. The animals set their well-being and maintenance as a prevailing priority of nutrient trafficking (homeostasis) at the expense of (growth) performance (homeorhesis). The lower feed intake in both RED and RED+RPMET led to a lower nutrient availability for metabolism, and hence, bulls grew only to that extent which could be realized with the difference between the total intake of nutrients and energy, and the requirements for maintenance. This concept is controlled at a higher endocrine organ level [In our previous study, bulls were allotted to three different treatment groups: a standard group (CON), which was designed to meet the requirements for German Simmental bulls for fattening at this stage of growth  and a dian level ,44 and tp > 0.05) dry matter and hence, energy and protein intake, which were, from a retrospective point of view, sufficient for their actually realized growth rates. Therefore, we could exclude both energy and protein intake as limiting factors for growth response in the RED+RPMET group. If methionine had been the first-limiting amino acid for growth, dietary protein quality  would have been improved. This means that more amino acids would have been utilized for protein deposition and this would have then resulted in a decrease in fat deposition in RED+RPMET compared to RED. Respective results can be found in [Considering the actually realized growth rates of both RED and RED+RPMET, the intake of pre-cecal digestible methionine met 101% of RED and 146% of RED+RPMET requirements, respectively. Both RED and RED+RPMET had equal  in cysteine glutathione disulfide and taurocholic acid, the conjugate of cholic acid and taurine, an important cellular antioxidant defender. The supplementation of RPMET (RED+RPMET) increased (p = 0.01) hepatic taurocholic acid concentrations as compared to the non-supplemented group  and reached a comparable level to the CON group (4.69). Taurocholic acid, a bile acid, is the conjugate of cholic acid and taurine. Taurine synthesis needs methionine [Additional methionine in the RED+RPMET group was directed toward anti-oxidant pathways, represented by an increase (thionine  via cystp < 0.01) in both CP-deficient groups (RED and RED+RPMET) as compared to the standard diet (CON). Interestingly, the addition of RPMET (RED+RPMET) failed to increase hepatic cystine concentrations. This would have been reasonable, since cystine is the oxidized form of cysteine, which is directly synthesized from methionine in the trans-sulfuration pathway [p < 0.01) the abundance of hepatic cysteine glutathione disulfide (4.96 in RED+RPMET vs. 4.63 in RED) and even exceeded the abundance in the standard group (4.79 in CON). Hepatic cystine was lower ( pathway . It may  pathway . InteresIn conclusion, our results show that growing German Simmental bulls face a dietary protein reduction with nutrient partitioning by setting maintenance and physiological equilibrium as the first priority in nutrient trafficking to ensure that all organs and tissues are interacting correctly. Additional methionine, which was not first limiting for growth under our feeding conditions , was dirSince our study was the first to elucidate the comprehensive metabolic role of methionine under such feeding conditions, further studies are required to sharpen the role of methionine in growing beef cattle. Further research should evaluate the ‘metabotype’, i.e., the combination of phenotype and metabolome , which oKnowledge on amino acid partitioning is particularly important to approach the ‘ideal protein concept’ for growing beef cattle. This is pivotal to increase the production efficiency of cattle genetic resources, thereby to drive further bio-economic circularity  and impr


Κ = 0.48).Prescribing errors represent a safety risk for hospitalized patients, especially in pediatrics. Computerized physician order entry (CPOE) might reduce prescribing errors, although its effect has not yet been thoroughly studied on pediatric general wards. This study investigated the impact of a CPOE on prescribing errors in children on general wards at the University Children’s Hospital Zurich. We performed medication reviews on a total of 1000 patients before and after the implementation of a CPOE. The CPOE included limited clinical decision support (CDS) such as drug-drug interaction check and checks for duplicates. Prescribing errors, their type according to the PCNE classification, their severity (adapted NCC MERP index), as well as the interrater reliability (Cohen’s kappa), were analyzed. Potentially harmful errors were significantly reduced from 18 errors/100 prescriptions (95% CI: 17–20) to 11 errors/100 prescriptions (95% CI: 9–12) after CPOE implementation. A large number of errors with low potential for harm  was reduced after the introduction of the CPOE, and consequently, the overall severity of potential harm increased post-CPOE. Despite general error rate reduction, medication reconciliation problems (PCNE error 8), such as drugs prescribed on paper as well as electronically, significantly increased after the introduction of the CPOE. The most common pediatric prescribing errors, the dosing errors (PCNE errors 3), were not altered on a statistically significant level after the introduction of the CPOE. Interrater reliability showed moderate agreement  was already in use before the implementation of the CPOE. Further investigations should focus on eliminating hybrid systems, interventions to increase the usability of the CPOE, and full integration of CDS tools such as automated dose checks into the CPOE.The online version contains supplementary material available at 10.1007/s00431-023-04894-5. Medication safety and the reduction of harm due to medication is an ongoing issue in health care. The WHO addresses this problem in the Third Global Patient Safety Challenge 2017, “Medication Without Harm.” . A meta-Computerized physician order entry (CPOE) seems to reduce prescribing errors –10, and On pediatric general wards with CPOE prescribing, errors range from 14.8 to 47.0 errors/100 prescriptions, whereas paper charts show a range from 4.1 to 58.1 errors/100 prescriptions . Dosing In general, studies about medication errors show a great heterogeneity in definitions and methods , 8, 13. The aim of this study was to investigate the impact of the introduction of a CPOE on prescribing errors in children in pediatric general wards. We studied the prevalence, type, and severity of prescribing errors with retrospective chart review and validated our findings by calculating the interrater reliability.This retrospective observational study was conducted at the University Children’s Hospital Zurich, which is a tertiary care center and the largest pediatric hospital in Switzerland (220 beds). Eligible for the study were children up to 18 years who were hospitalized in 3 medical or 3 surgical wards. A total of 1000 patients, 500 from each study period, were randomly selected from 1688 (2018) and 1608 (2019) eligible patients, respectively, consisting of 250 medical and 250 surgical cases each. Patients with no prescribed medication  were not eligible. To control for seasonal effects, periods in identical months were included in the sampling frame: October until December 2018 (pre-CPOE) and October until December 2019 (post-CPOE).Until 2019 drugs were prescribed either manually on paper charts or electronically on semi-structured charts. CGM Clinical by CompuGroup Medical Inc. was used as a hospital information system. The pediatrics-specific CPOE tool by CompuGroup Medical Inc. was further developed from the preexisting CPOE tool for adults in cooperation with members of the University Children’s Hospital Zurich.The drug master data were provided by HospINDEX (HCI Solutions Ltd.) and validated by the hospital pharmacy. Drugs from the hospital formulary were marked and provided with appropriate routes of administration so that only these routes of administration could be selected by the prescribing physician in the ordering process. In case of drug shortages, the appropriate substitute was proposed to the prescriber.The CPOE contained some limited clinical decision support (CDS): an automated drug-drug interaction check and a duplication check based on the data of Pharmavista by HCI Solutions Ltd. , which wIn March 2019, the CPOE was implemented on all general wards. Implementation was accompanied by user education. Thereafter, most of the medication was prescribed electronically with the CPOE. There were some exceptions that could not be prescribed with the tool for technical reasons, such as patient-controlled analgesia and others, leading to hybrid drug prescription system to a small degree.Prescribers at the University Children’s Hospital Zurich had access to several guidelines and databases. They were requested to follow the dosing guidelines provided by the web application CDS by PEDeDose (PEDeus Ltd.), which was described by Higi et al. . These dA full medication review was conducted for all included patients, whereby the rater had access to all patient data. An adapted version of the medication appropriateness index  was usedAll prescriptions within the first 24 h of admission to the ward were included. For patients who were hospitalized for elective surgery, prescriptions were included in the 24 h after surgery and return to the ward from PICU. Excluded medications were parenteral nutrition, lipids, any blood cell transfusions, insulin, solutions for dialysis, solutions for fluid management such as normal saline solution, dextrose 5%, dextrose 5% in normal saline solution, or acetated ringers. Electrolytes were included, even if they were added to parenteral nutrition.As proposed by Dean et al.  the defiPrescribing errors were classified into different types of errors according to the well-established PCNE Classification V 9.1, German Version , 22. As To increase the clinical relevance of our findings, we also included the severity of harm due to errors, even though the retrospective rating of the potential harm is likely to be more subjective than the rating of actual harm. , 15. To All patients were reviewed by the first rater (AS), and a random sample of 5% of all patients was additionally reviewed by a second rater (MP). Both raters were pharmacists with several years of experience in pediatric clinical pharmacy. A procedural manual was created to ensure that both raters approached the review in a similar way and that consistent data could be captured, as suggested by Vassar et al. . Both raThe study database was built with Microsoft SQL Server 2019 Master Data Services. The personal data from all included patients were automatically exported from the hospital information system into the database.® SPSS® Statistics Version 27. The sample size of 1000 patients was estimated according to a previous study by Glanzmann et al. [T-test, chi-square test, or Mann–Whitney test. Interrater reliability was calculated with Cohen’s kappa.Statistical analyses were conducted with RStudio 2022.02.1 and IBMp = 0.005, t-test).Pre-CPOE and post-CPOE patients did not differ in their demographic characteristics Table , except p < 0.001).A total of 5022 drug prescriptions for 1000 patients were analyzed: 2299 drug prescriptions pre-CPOE and 2723 post-CPOE. Patients were prescribed more drugs post-CPOE than pre-CPOE .Handwritten prescriptions were significantly reduced by the CPOE . This means that 69.2% (95% CI: 67.4–71.0) of all prescriptions pre-CPOE contained at least one error vs. only 22.8% (95% CI: 21.2–24.5) of all prescriptions post-CPOE, which implies a significant reduction of errors (p < 0.001). In 100 admissions, 360 errors could be found pre-CPOE vs. 137 in post-CPOE (p < 0.001).A total of 2485 errors for all prescriptions was found, of which 1802 errors occurred pre-CPOE and 683 errors post-CPOE. Consequently, 78.4 errors per 100 prescriptions (95% CI: 76.2–80.6) were found before the introduction of the CPOE vs. 25.1 errors per 100 prescriptions (95% CI: 23.0–27.1) after introduction of the CPOE  was in PCNE primary domain 5 (dispensing) .The overall severity of harm increased significantly after the introduction of the CPOE. Mean rank pre-CPOE was 1175 vs. 1423 post-CPOE : 76.4% pre-CPOE and 58% post-CPOE (Table p < 0.001). Therefore, 16.8% of all prescriptions before CPOE contained at least one error (95% CI: 15.5–18.2), whereas the error rate after CPOE was only 9.8% (95% CI: 8.6–11.1) (p < 0.001). The rate of errors per 100 admissions was significantly reduced from 84 errors/100 admissions pre-CPOE to 57 errors/100 admissions post-CPOE (p = 0.001).After exclusion of errors with severity A–D, we counted a remaining total of 711 errors with potential severity of harm, E–I, which were 424 errors pre-CPOE and 287 errors post-CPOE. The overall error rate of 18.4 errors/100 prescriptions (95% CI: 16.8–19.9) before CPOE was reduced to 10.5 errors/100 prescriptions (95% CI: 9.1–12.0) after introduction of the CPOE, which was a significant reduction of the error rate (p < 0.001), and type 1 errors (drug selection) decreased from 3.0 to 2.0 errors/100 prescriptions (p = 0.031).As Table p = 0.010). Type 8 errors were coded when two valid prescriptions for the same patient and time were found in different media, e.g., one CPOE prescription and one prescription on a paper chart. The doubled prescriptions contained, in some cases, the same information but sometimes slightly different dosages or instructions.The third domain that showed a significant change from pre- to post-CPOE was PCNE 8 “patient transfer related.” These errors increased significantly from 0.2 errors/100 prescriptions to 0.6 errors/100 prescriptions (p = 0.555). The PCNE causes are also displayed in Table There were no statistically significant differences from pre- to post-CPOE in the other primary domains. In particular, dosing errors (type 3) showed no significant change . A total of 20.9% of handwritten prescriptions contained at least one error, while only 14.2% of electronically written orders contained an error.Pre-CPOE, we compared the error rates in handwritten and electronically written prescriptions. Handwritten prescriptions showed a significantly higher rate of errors (22.6 errors/100 prescriptions) than electronically written prescriptions (15.7 errors/100 prescriptions)  was calculated.Cohen’s Kappa for the agreement on whether or not an error occurred in a prescription was 0.476. This implies a moderate interrater agreement . The agrAfter the implementation of the CPOE, patients were prescribed more drugs than before (5.5 vs. 4.6). The fact that many adjustments to a CPOE prescription, such as an adaption of the dose, resulted in a new prescription line may have influenced this finding.The overall error rate decreased significantly after implementation of the CPOE. This finding complies with the results of a recent systematic review in pediatrics by Koeck et al. , indicatThe rate of potentially harmful errors (NCC MERP E-I) of 18.4% (95% CI 16.8–19.9) before introduction of the CPOE was higher than the error rate of 14%. Glanzmann et al.  found 9 p < 0.001). This is consistent with the findings of Jungreithmayr et al. [The CPOE especially improves the quality of prescriptions, which can be seen in the largest reduction of potentially harmful errors (NCC MERP E-I) in PCNE errors 5.2 (“lacking or wrong information”) from 8.7 to 1.9 errors/100 prescriptions  and especially cause 1.4 “inappropriate duplication of therapeutic group or active ingredient” were significantly reduced post-CPOE. This could be contributed to the duplication-check CDS.Interestingly, cause 1.3 concerning drug-drug interactions did not decrease, even though one CDS tool (Pharmavista) offered an automatic drug-drug interaction check. This could be attributed to either alert fatigue  or poor Nevertheless, a new hybrid-systems-related error did occur, as medication reconciliation problems (PCNE 8) increased significantly with remaining paper prescriptions for special medication. This type of error might be prevented through the integration of all prescriptions into the CPOE and the complete elimination of prescriptions on paper.The CPOE had no effect on dosing errors. This finding might be explained by the fact that the web application CDS covering dosing recommendations (PEDeDose) was already in use before implementation of the CPOE, and there was no automated dose check to validate the prescriptions. The lacking effect of CPOE on dosing errors was also seen by Roumeliotis et al. . Dosing The fact that handwritten prescriptions contained more errors than electronic prescriptions is a plausible finding, as CPOE leads to standardized, legible, and complete prescriptions .The errors that occurred post-CPOE were more severe than pre-CPOE. This implies that a large number of minor-severity errors like “missing information” (PNCE 5.2) and others no longer occurred after the introduction of the CPOE. It also indicates that the introduction of the CPOE was not able to reduce the severity of harm but only the rate of errors.Due to the retrospective nature of our study, we had to interpret previously recorded data on patients’ history and their prescriptions. This could have led to the conclusion that an error had occurred, although there was a plausible reason for the deviation from the norm, which was however not recorded in the patient’s charts. For this reason, the retrospective nature of our study imposes a limitation not only on the rate of errors but especially on the interpretation of the severity of harm.Furthermore, not optimal interrater reliability imposes another limitation to our study. Even though a procedure manual existed, there was only limited training and coordination between the two raters. One rater might have tried to capture errors comprehensively, whereas the other had a more pragmatic way of assessment of the prescriptions. Rater 1 reviewed all 1000 patients and therefore had more routine in the procedure, while rater 2 only reviewed 50 patients.In conclusion, our findings imply a positive impact of the CPOE on patient safety by reducing the prevalence of prescribing errors. Especially the high number of errors with low harming potential (NCC MERP A–D) was reduced, but also potentially harmful errors were significantly reduced.Hybrid systems of CPOE and paper charts carry a risk for errors and should therefore be eliminated. Future research might focus on interventions to increase the usability of the CPOE and on the full integration of CDS tools, for example, dosing support and notably automated dose check into the CPOE.Supplementary file1 Adapted MAI (DOCX 15 KB)Supplementary file2 Rating of dosing errors (DOCX 19 KB)Below is the link to the electronic supplementary material.


BackgroundMinimally invasive liver resections for metastatic colorectal cancer have been increasingly performed all over the world with promising results. We planned the current study to review our experience on this matter and compare short- and long-term outcomes of laparoscopic liver resection (LLR) and open liver resection (OLR) in patients with colorectal cancer liver metastasis (CRLM).Materials and methodsThis is a single-center retrospective analysis of patients with CRLM who underwent laparoscopic (n=86) and open (n=96) surgical treatment for metastatic liver lesions between March 2016 and November 2022. Tumor characteristics, intra- and postoperative results, overall survival (OS), and disease-free survival (DFS) were analyzed and compared.ResultsLLR was associated with significantly shorter surgery duration . There was no significant difference in blood loss between the two groups . Additionally, the laparoscopic approach was associated with significantly shorter hospital stays . The rate of major complications (Clavien-Dindo classification ≥ 3) was lower in the LLR group . There was no mortality in the LLR group, and in the OLR group, one lethal case was induced by mesenteric thrombosis on the fifth postoperative day. We did not find a statistically significant difference in the OS rate between the two groups at one, three, and five years: 97.3%, 74.7%, and 43.4%, respectively, in the OLR group and 95.1%, 70.3%, and 49.5%, respectively, in the LLR group (p=0.53). DFS at one, three, and five years were 88.7%, 52.3%, and 25.5%, respectively, in the LLR group and 71.9%, 53.1%, and 19.3%, respectively, in the OLR group (p=0.66).ConclusionsThis study showed that laparoscopic liver surgery is a safe and effective method of CRLM treatment in our center. LLR was associated with a decrease in major morbidity, shorter surgery duration, and reduced postoperative hospital stay. Minimally invasive liver resections showed similar oncological outcomes to the open approach in terms of overall and disease-free survival. In 2020, colorectal cancer was diagnosed in more than 1.9 million people, ranking third in terms of incidence and second in terms of cancer-related mortality, accounting for 9.4% of lethal cases . The maiScientific evidence on the feasibility of minimally invasive techniques in the field of liver surgery is confined to retrospective series, meta-analysis, and results of randomized controlled trials (RCTs), such as the OSLO-COMET and LapOpHuva -7. The oPatient selectionWe performed a retrospective analysis of colorectal cancer patients with liver metastasis (CRLM) who received laparoscopic (n=85) and open (n=96) surgical treatment for metastatic liver lesions between March 2016 and November 2022 at the National Cancer Institute.The inclusion criterion was a curative intended resection directed toward the removal of all radiologically evident liver lesions. In total, 181 CRLM patients with morphologically confirmed adenocarcinoma of the colon or rectum and ≥1 liver metastasis were included in the study. We excluded patients < 18 years old, those who carried more than five resectable lung metastases, and/or unresectable peritoneal carcinomatosis.The preoperative investigations were conducted according to international standards: blood tests, contrast-enhanced CT, and/or MRI. All patients were discussed at the multidisciplinary board meeting. Liver anatomy and resection were classified according to The Brisbane 2000 Terminology, based on Couinaud’s segments .Short- and long-term outcomesThe clinical parameters examined were age, gender, American Society of Anesthesiologists (ASA) score, body mass index (BMI), and CRLM characteristics. Surgical parameters such as the type of resection, operation time, conversion rate, and blood loss were collected. Primary outcomes were assessed by postoperative morbidity, mortality, and length of hospital stay. Major complications were defined either as requiring intensive care unit stay, treatment by an interventional radiologist, or reoperation . Long-teSurgical techniqueSurgical techniques included anatomical-oriented, parenchymal-sparing open liver resection (OLR) and laparoscopic liver resection (LLR), and radiofrequency ablation (RFA) for deeply located tumors ≤ 2 cm. Major hepatectomy was defined as the resection of three or more contiguous liver segments. All surgeries were performed under general anesthesia. To locate the tumor and guide the resection plane according to anatomical landmarks, we used intraoperative ultrasonography. The patient’s position and operative setup for laparoscopic resections were described in detail elsewhere ,11. TrocStatistics2) (with Yates’ correction or not) and Fisher’s exact test. P-values of <0.05 were considered statistically significant. Survival was evaluated via the Kaplan-Meier method and compared using the log-rank test.For nonparametric data, to define the normality of distribution, the Kolmogorov-Smirnov test was used. Continuous variables were expressed as median or mean and range interval depending on the normality of distribution. To compare normally distributed data, the Mann-Whitney U test was used. For data with skewed distribution, Student’s t-test was applied. Categorical variables were reported as number (n) and percentage (%) and compared using the chi-squared test  patients underwent liver resection via laparoscopic approach, and 96 (53%) underwent open liver resections (OLR). The primary characteristics of patients are presented in Table Operative and postoperative dataFigure There was no difference in the amount of resected liver segments between the two groups (p=0.07). In the OLR group, 8.3% (8/96) of patients underwent major liver resections, and in the LLR group, it was 3.5% (3/85). In the laparoscopic liver resection group, we performed radiofrequency ablation (RFA) in 8.2% (7/85) of patients . In the LLR group, there were five conversions (5.9%) due to intra-abdominal adhesions after previous colon and/or liver resections and the inability to safely perform adhesiolysis via a laparoscopic approach.As shown in Table The median survival was 36.2 months and 38 months in the LLR and OLR groups, respectively. As Figure This study compares the results of laparoscopic and open liver resections in patients with colorectal cancer liver metastasis who were treated in the settings of a Ukrainian state center. We report successful short- and long-term outcomes of laparoscopic liver resection in patients with CRLM.A number of retrospective studies showed that laparoscopic liver resections are more favorable in terms of postoperative complication rate -15. OtheAs to the long-term outcomes, multiple studies have proven that the laparoscopic approach does not impair overall survival ,22-24. WStarting from the consensus conference held in Louisville, laparoscopic resections of posterosuperior liver segments (PSS) have been considered “major” from a technical point of view and were suggested to be performed at expert centers only . Teo et Implementation of a highly specialized surgical project such as a laparoscopic liver resection program requires significant initial and sustained efforts . AnalysiThe present study has a number of limitations. Firstly, retrospective studies are prone to selection bias with regard to operative methods. Secondly, conclusions should be carefully withdrawn due to a rather small cohort of patients. Thirdly, since the data was collected from a single institution, there is a place for center-related bias. Additionally, the beginning of the study period was associated with a learning curve that could influence the results.Surgical resection remains the most radical treatment option for patients with colorectal cancer liver metastasis. A minimally invasive approach can be an advantageous strategy for such patients. We found that LLRs are associated with better short-term outcomes: decreased major morbidity, shorter surgery duration, and significantly lower postoperative hospital stay. As for oncological outcomes, LLRs are equivalent to open resections in terms of overall and disease-free survival. Therefore, laparoscopic liver resections are a feasible method of surgical treatment, safely adapted to the reality of our institution.