Lithocholic Acid Research Articles

Bifidobacterium animalis subsp. lactis F1-7 Alleviates Lipid Accumulation in Atherosclerotic Mice via Modulating Bile Acid Metabolites to Downregulate Intestinal FXR.

The dysfunction of intestinal microbiota and bile acid metabolism is related to the pathogenesis of atherosclerosis. This study we explored the mechanism of Bifidobacterium animalis subsp. lactis F1-7 (Bif. animalis F1-7), improving atherosclerosis by regulating the bile acid metabolism and intestinal microbiota in the ApoE-/- mice. The Bif. animalis F1-7 effectively reduced aortic plaque accumulation and improved the serum and liver lipid levels in atherosclerotic mice. The untargeted metabolomics revealed that Bif. animalis F1-7 reduced the glycine-conjugated bile acids and the levels of differential metabolite lithocholic acid (LCA) significantly. Downregulation of LCA decreased the intestinal levels of the farnesoid X-activated receptor (FXR) and regulated the bile acid metabolism through the FXR/FGF15/CYP7A1 pathway. Furthermore, the 16srRNA gene sequencing analysis revealed that structural changes in intestinal microbiota with an increase in the abundance of Bifidobacterium, Lactobacillus, Faecalibaculum, Desulfovibrio, and a decrease in Dubosiella, Clostridium_sensu_stricto_1, and Turicibacter following the Bif. animalis F1-7 intervention. Correlation analysis showed that the changes in intestinal microbiota mentioned above were significantly correlated with bile acid metabolism in atherosclerotic mice. In conclusion, this study sheds light on the mechanisms by which Bif. animalis F1-7 regulates atherosclerosis.

Antimicrobial activity and properties of de novo design of short synthetic lipopeptides

The aim of this article is to introduce the topic of newly designed peptides as well as their biological activity. We designed nine encoded peptides composed of six amino acids. All these peptides were synthesized with C-terminal amidation. To investigate the importance of increased hydrophobicity at the amino end of the peptides, all of them were subsequently synthesized with palmitic or lithocholic acid at the N-terminus. Antimicrobial activity was tested on Gram-positive and Gram-negative bacteria and fungi. Cytotoxicity was measured on HepG2 and HEK 293 T cell cultures. Peptides bearing a hydrophobic group exhibited the best antimicrobial activity. Lipopeptides with palmitic or lithocholic acid (PAL or LCA peptides) at the N-terminus and with C-terminal amidation were highly active against Gram-positive bacteria, especially against strains of Staphylococcus aureus and Candida tropicalis. The LCA peptide SHP 1.3 with the sequence LCA-LVKRAG-NH2, had high efficiency on HepG2 human liver hepatocellular carcinoma cells (97%).

P369 Altered fecal bile acid composition in active Ulcerative Colitis

Abstract Background A consistent finding in inflammatory bowel disease (IBD) is an altered composition of fecal bile acids, with an increase in primary bile acids and a decrease in secondary bile acids. It is less clear, whether fecal bile acids could prove to be biomarkers for IBD diagnosis and disease activity. The study aimed to determine correlations between eighteen fecal bile acid species and IBD entity as well as disease severity. Methods Eighteen fecal bile acid species were quantified from stool samples of 62 IBD patients and 17 controls using LC-MS/MS and stable isotope dilution. Bile acid levels were normalized to the dry weight of the fecal homogenates. For calculations, normalized bile acid concentrations and ratios of individual bile acid species to total bile acid levels were used. The p-values were corrected for multiple comparisons. Results In accordance with previous data, we showed that patients with IBD had more primary and less secondary bile acids in their stool compared to healthy controls, with greater differences when comparing healthy controls with ulcerative colitis (UC) than with Crohn´s disease (CD). In CD patients’ fecal calprotectin showed no correlations with any of the bile acids. In UC negative correlations between fecal calprotectin levels and two secondary bile acids, glycine conjugated lithocholic acid (GLCA) and hyodeoxycholic acid (HDCA), and thus total levels of secondary bile acids, existed. Comparing patients with low calprotectin levels (<50 µg/g) and patients with high calprotectin levels (>500 µg/g) in UC revealed that the latter group had lower GLCA and HDCA. Fecal bile acid levels of CD patients did not change with higher calprotectin. Moreover, serum C-reactive protein negatively correlated with the secondary bile acids HDCA, ursodeoxycholic acid, lithocholic acid, taurine conjugated lithocholic acid, deoxycholic acid and its taurine conjugated form in UC but not CD patients. Conclusion To distinguish between IBD subtypes, it may be helpful to note that impaired fecal bile acid homeostasis is specific to patients with active UC.

Impact of lithocholic acid on the osteogenic and adipogenic differentiation balance of bone marrow mesenchymal stem cells

To Investigate the effects of lithocholic acid (LCA) on the balance between osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). Twelve 10-week-old SPF C57BL/6J female mice were randomly divided into an experimental group (undergoing bilateral ovariectomy) and a control group (only removing the same volume of adipose tissue around the ovaries), with 6 mice in each group. The body mass was measured every week after operation. After 4 weeks post-surgery, the weight of mouse uterus was measured, femur specimens of the mice were taken for micro-CT scanning and three-dimensional reconstruction to analyze changes in bone mass. Tibia specimens were taken for HE staining to calculate the number and area of bone marrow adipocytes in the marrow cavity area. ELISA was used to detect the expression of bone turnover markers in the serum. Liver samples were subjected to real-time fluorescence quantitative PCR (RT-qPCR) to detect the expression of key genes related to bile acid metabolism, including cyp7a1, cyp7b1, cyp8b1, and cyp27a1. BMSCs were isolated by centrifugation from 2 C57BL/6J female mice (10-week-old). The third-generation cells were exposed to 0, 1, 10, and 100 μmol/L LCA, following which cell viability was evaluated using the cell counting kit 8 assay. Subsequently, alkaline phosphatase (ALP) staining and oil red O staining were conducted after 7 days of osteogenic and adipogenic induction. RT-qPCR was employed to analyze the expressions of osteogenic-related genes, namely ALP, Runt-related transcription factor 2 (Runx2), and osteocalcin (OCN), as well as adipogenic-related genes including Adiponectin (Adipoq), fatty acid binding protein 4 (FABP4), and peroxisome proliferator-activated receptor γ (PPARγ). Compared with the control group, the body mass of the mice in the experimental group increased, the uterus atrophied, the bone mass decreased, the bone marrow fat expanded, and the bone metabolism showed a high bone turnover state. RT-qPCR showed that the expressions of cyp7a1, cyp8b1, and cyp27a1, which were related to the key enzymes of bile acid metabolism in the liver, decreased significantly ( P<0.05), while the expression of cyp7b1 had no significant difference ( P>0.05). Intervention with LCA at concentrations of 1, 10, and 100 μmol/L did not demonstrate any apparent toxic effects on BMSCs. Furthermore, LCA inhibited the expressions of osteogenic-related genes (ALP, Runx2, and OCN) in a dose-dependent manner, resulting in a reduction in ALP staining positive area. Concurrently, LCA promoted the expressions of adipogenic-related genes (Adipoq, FABP4, and PPARγ), and an increase in oil red O staining positive area. After menopause, the metabolism of bile acids is altered, and secondary bile acid LCA interferes with the balance of osteogenic and adipogenic differentiation of BMSCs, thereby affecting bone remodelling.

Gender-Specific Bile Acid Profiles in Non-Alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) is increasing worldwide. A main cause is the obesogenic, so-called Western lifestyle. NAFLD follows a long, unperceived course, and ends potentially fatally. Early diagnosis of aggressive subtypes saves lives. So far, non-invasive means of detection are limited. A better understanding of the pathogenic interplay among insulin resistance, immune inflammation, microbiome, and genetic background is important. Metabolomics may give insight into these interlaced processes. In this study, we measured bile acids (BA) in the plasma of adult NAFLD and alcohol-associated liver disease (ALD) patients and healthy controls with targeted mass spectrometry. We focused on gender-related bile acid production pathology in NAFLD and ALD. Compared to healthy controls, women with NAFLD had significantly higher concentrations of total BA, total primary BA, total cholic (CA), total chenodeoxycholic (CDCA), total glycine-conjugated, and total non-12-a-OH BA. Concerning subtypes, glycocholic (GCA) and glycochenodeoxycholic (GCDCA), BA were elevated in women with NAFLD. In contrast, men with NAFLD had no significantly altered total BA fractions. However, the subtypes GCA, glycodeoxycholic (GDCA), glycolithocholic (GLCA), lithocholic (LCA), taurolithocholic (TLCA), and tauroursodeoxycholic acid (TUDCA) were elevated, while CA was significantly decreased. In NAFLD, except ursodeoxycholic acid (UDC), all total BA correlated significantly positively in both sexes with the ELF score, while in ALD, only males showed significant correlations exceptive for total UDC BA. In NAFLD, total BA, total primary BA, total secondary BA, total free secondary BA, total CA, total CDCA, total taurine conjugated, total glycine conjugated, total 12-a-OH, and total non-12-a-OH were significantly higher in cases of a high enhanced liver fibrosis (ELF) score above 9.8. In ALD, total UDC was additionally elevated. Between NAFLD with and without NASH, we found no significant differences. Our data show gender-specific bile acid profiles in NAFLD and markedly different BA patterns in ALD. Women with NAFLD had more severe cholestasis. Men may better compensate fat storage-driven bile acid dynamics, indicated by higher levels of taurine-conjugated BA, which associate with beneficial metabolic functions.

Chicoric Acid Effectively Mitigated Dextran Sulfate Sodium (DSS)-Induced Colitis in BALB/c Mice by Modulating the Gut Microbiota and Fecal Metabolites.

Chicoric acid (CA) has been reported to exhibit biological activities; it remains unclear, however, whether CA could regulate colitis via modulation of the gut microbiota and metabolites. This study aimed to assess CA's impact on dextran sulfate sodium (DSS)-induced colitis, the gut microbiota, and metabolites. Mice were induced with 2.5% DSS to develop colitis over a 7-day period. CA was administered intragastrically one week prior to DSS treatment and continued for 14 days. The microbial composition in the stool was determined using 16S rRNA sequencing, while non-targeted metabolomics was employed to analyze the metabolic profiles of each mouse group. The results show that CA effectively alleviated colitis, as evidenced by an increased colon length, lowered disease activity index (DAI) and histological scores, and decreased tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) expression levels. CA intervention restored the structure of gut microbiota. Specifically, it decreased the abundance of Bacteroidetes and Cyanobacteria at the phylum level and Bacteroides, Rosiarcus, and unclassified Xanthobacteraceae at the genus level, and increased the abundance of unclassified Lachnospiraceae at the genus level. Metabolomic analysis revealed that CA supplementation reversed the up-regulation of asymmetric dimethylarginine, N-glycolylneuraminic acid, and N-acetylneuraminic acid, as well as the down-regulation of phloroglucinol, thiamine, 4-methyl-5-thiazoleethanol, lithocholic acid, and oxymatrine induced by DSS. Our current research provides scientific evidence for developing CA into an anti-colitis functional food ingredient. Further clinical trials are warranted to elucidate the efficacy and mechanism of CA in treating human inflammatory bowel disease (IBD).

Bile acid-containing lipid nanoparticles enhance extrahepatic mRNA delivery.

Lipid nanoparticles (LNPs) have emerged as a viable, clinically-validated platform for the delivery of mRNA therapeutics. LNPs have been utilized as mRNA delivery systems for applications including vaccines, gene therapy, and cancer immunotherapy. However, LNPs, which are typically composed of ionizable lipids, cholesterol, helper lipids, and lipid-anchored polyethylene glycol, often traffic to the liver which limits the therapeutic potential of the platform. Several approaches have been proposed to resolve this tropism such as post-synthesis surface modification or the addition of synthetic cationic lipids. Methods: Here, we present a strategy for achieving extrahepatic delivery of mRNA involving the incorporation of bile acids, a naturally-occurring class of cholesterol analogs, during LNP synthesis. We synthesized a series of bile acid-containing C14-4 LNPs by replacing cholesterol with bile acids (cholic acid, chenodeoxycholic acid, deoxycholic acid, or lithocholic acid) at various ratios. Results: Bile acid-containing LNPs (BA-LNPs) were able to reduce delivery to liver cells in vitro and improve delivery in a variety of other cell types, including T cells, B cells, and epithelial cells. Our subsequent in vivo screening of selected LNP candidates injected intraperitoneally or intravenously identified a highly spleen tropic BA-LNP: CA-100, a four-component LNP containing cholic acid and no cholesterol. These screens also identified BA-LNP candidates demonstrating promise for other mRNA therapeutic applications such as for gastrointestinal or immune cell delivery. We further found that the substitution of cholic acid for cholesterol in an LNP formulation utilizing a different ionizable lipid, C12-200, also shifted mRNA delivery from the liver to the spleen, suggesting that this cholic acid replacement strategy may be generalizable. Conclusion: These results demonstrate the potential of a four-component BA-LNP formulation, CA-100, for extrahepatic mRNA delivery that could potentially be utilized for a range of therapeutic and vaccine applications.

Chronic arsenic exposure-provoked biotoxicity involved in liver-microbiota-gut axis disruption in chickens based on multi-omics technologies

IntroductionArsenic has been ranked as the most hazardous substance by the U.S. Agency for Toxic Substances and Disease Registry. Environmental arsenic exposure-evoked health risks have become a vital public health concern worldwide owing to the widespread existence of arsenic. Multi-omics is a revolutionary technique to data analysis providing an integrated view of bioinformation for comprehensively and systematically understanding the elaborate mechanism of diseases. ObjectivesThis study aimed at uncovering the potential contribution of liver-microbiota-gut axis in chronic inorganic arsenic exposure-triggered biotoxicity in chickens based on multi-omics technologies. MethodsForty Hy-Line W-80 laying hens were chronically exposed to sodium arsenite with a dose-dependent manner (administered with drinking water containing 10, 20, or 30 mg/L arsenic, respectively) for 42 d, followed by transcriptomics, serum non-targeted metabolome, and 16S ribosomal RNA gene sequencing accordingly. ResultsArsenic intervention induced a serious of chicken liver dysfunction, especially severe liver fibrosis, simultaneously altered ileal microbiota populations, impaired chicken intestinal barrier, further drove enterogenous lipopolysaccharides translocation via portal vein circulation aggravating liver damage. Furtherly, the injured liver disturbed bile acids (BAs) homoeostasis through strongly up-regulating the BAs synthesis key rate-limiting enzyme CYP7A1, inducing excessive serum total BAs accumulation, accompanied by the massive synthesis of primary BA—chenodeoxycholic acid. Moreover, the concentrations of secondary BAs—ursodeoxycholic acid and lithocholic acid were markedly repressed, which might involve in the repressed dehydroxylation of Ruminococcaceae and Lachnospiraceae families. Abnormal BAs metabolism in turn promoted intestinal injury, ultimately perpetuating pernicious circle in chickens. Notably, obvious depletion in the abundance of four profitable microbiota, Christensenellaceae, Ruminococcaceae, Muribaculaceae, and Faecalibacterium, were correlated tightly with this hepato-intestinal circulation process in chickens exposed to arsenic. ConclusionOur study demonstrates that chronic inorganic arsenic exposure evokes liver-microbiota-gut axis disruption in chickens and establishes a scientific basis for evaluating health risk induced by environmental pollutant arsenic.

Quantitative bile acid profiling in healthy adult dogs and pups from serum, plasma, urine, and feces using LC-MS/MS.

Synthesis and secretion of bile acids (BA) is a key physiological function of the liver. In pathological conditions like portosystemic shunt, hepatic insufficiency, hepatitis, or cirrhosis BA metabolism and secretion are disturbed. Quantification of total serum BA is an established diagnostic method to assess the general liver function and allows early detection of abnormalities, liver disease progression and guidance of treatment decisions. To date, data on comparative BA profiles in dogs are limited. However, BA profiles might be even better diagnostic parameters than total BA concentrations. On this background, the present study analyzed and compared individual BA profiles in serum, plasma, urine, and feces of 10 healthy pups and 40 adult healthy dogs using ultra-high performance liquid chromatography coupled to electrospray ionization mass spectrometry. Sample preparation was performed by solid-phase extraction for serum, plasma, and urine samples or by protein precipitation with methanol for the feces samples. For each dog, 22 different BA, including unconjugated BA and their glycine and taurine conjugates, were analyzed. In general, there was a great interindividual variation for the concentrations of single BA, mostly exemplified by the fact that cholic acid (CA) was by far the most prominent BA in blood and urine samples of some of the dogs (adults and pups), while in others, CA was under the detection limit. There were no significant age-related differences in the BA profiles, but pups showed generally lower absolute BA concentrations in serum, plasma, and urine. Taurine-conjugated BA were predominant in the serum and plasma of both pups (68%) and adults (74-75%), while unconjugated BA were predominant in the urine and feces of pups (64 and 95%, respectively) and adults (68 and 99%, respectively). The primary BA chenodeoxycholic acid and taurocholic acid and the secondary BA deoxycholic acid and lithocholic acid were the most robust analytes for potential diagnostic purpose. In conclusion, this study reports simultaneous BA profiling in dog serum, plasma, urine, and feces and provides valuable diagnostic data for subsequent clinical studies in dogs with different kinds of liver diseases.

Dietary nobiletin regulated cefuroxime- and levofloxacin-associated "gut microbiota-metabolism" imbalance and intestinal barrier dysfunction in mice.

Nobiletin (NOB) exhibits significant biological activities and may be a potential dietary treatment for antibiotic-associated gut dysbiosis. In this study, mice were gavaged with 0.2 mL day-1 of 12.5 g L-1 cefuroxime (LFX) and 10 g L-1 levofloxacin (LVX) for a duration of 10 days, accompanied by 0.05% NOB to investigate the regulatory effect and potential mechanisms of NOB on antibiotic-induced intestinal microbiota disorder and intestinal barrier dysfunction. Our results indicated that dietary NOB improved the pathology of intestinal epithelial cells and the intestinal permeability by upregulating the expression of intestinal tight junction proteins (TJs) and the number of goblet cells. Furthermore, dietary NOB reduced the levels of serum lipopolysaccharide (LPS) and pro-inflammatory factors (TNF-α and IL-1β), thereby facilitating the restoration of the intestinal mucosal barrier. Additionally, dietary NOB increased the abundance of beneficial bacteria f_Lachnospiraceae and regulated the metabolic disorders of short-chain fatty acids (SCFAs) and bile acids (BAs). Notably, NOB supplementation resulted in elevated levels of butyric acid and lithocholic acid (LCA), which contributed to the repair of the intestinal mucosal barrier function and the maintenance of intestinal homeostasis. Collectively, our results propose a healthy dietary strategy for the prevention or mitigation of antibiotic-associated gut dysbiosis by dietary NOB.

Bifidobacterium longum GL001 alleviates rat intestinal ischemia-reperfusion injury by modulating gut microbiota composition and intestinal tissue metabolism.

Intestinal ischemia-reperfusion (IIR) injury leads to inflammation and oxidative stress, resulting in intestinal barrier damage. Probiotics, due to their anti-inflammatory and antioxidant properties, are considered for potential intervention to protect the intestinal barrier during IIR injury. Bifidobacterium longum, a recognized probiotic, has targeted effects on IIR injury, but its mechanisms of action are not yet understood. To investigate the mechanism of Bifidobacterium longum intervention in IIR injury, we conducted a study using a rat IIR injury model. The results showed that Bifidobacterium longum could alleviate inflammation and oxidative stress induced by IIR injury by suppressing the NF-κB inflammatory pathway and activating the Keap1/Nrf2 signaling pathway. Bifidobacterium longum GL001 also increased the abundance of the gut microbiota such as Oscillospira, Ouminococcus, Corynebacterium, Lactobacillus, and Akkermansia, while decreasing the abundance of Allobaculum, [Prevotella], Bacteroidaceae, Bacteroides, Shigella, and Helicobacter. In addition, Bifidobacterium longum GL001 reversed the changes in amino acids and bile acids induced by IIR injury and reduced the levels of DL-cysteine, an oxidative stress marker, in intestinal tissue. Spearman correlation analysis showed that L-cystine was positively correlated with Lactobacillus and negatively correlated with Shigella, while DL-proline was positively correlated with Akkermansia. Moreover, bile acids, cholic acid and lithocholic acid, were negatively correlated with Lactobacillus and positively correlated with Shigella. Therefore, Bifidobacterium longum GL001 may alleviate IIR injury by regulating the gut microbiota to modulate intestinal lipid peroxidation and bile acid metabolism.

Engineered nanomicelles targeting proliferation and angiogenesis inhibit tumour progression by impairing the synthesis of ceramide-1-phosphate.

Tumour cells secrete various proangiogenic factors like VEGF, PDGF, and EGF that result in the formation of highly vascularized tumours with an immunosuppressive tumour microenvironment. As tumour growth and metastasis are highly dependent on angiogenesis, targeting tumour vasculature along with rapidly dividing tumour cells is a potential approach for cancer treatment. Here, we specifically engineered sub-100 sized nanomicelles (DTX-CA4 NMs) targeting proliferation and angiogenesis using an esterase-sensitive phosphocholine-tethered docetaxel conjugate of lithocholic acid (LCA) (PC-LCA-DTX) and a poly(ethylene glycol) (PEG) derivative of an LCA-combretastatin A4 conjugate (PEG-LCA-CA4). DTX-CA4 NMs effectively inhibit the tumour growth in syngeneic (CT26) and xenograft (HCT116) colorectal cancer models, inhibit tumour recurrence, and enhance the percentage survival in comparison with individual drug-loaded NMs. DTX-CA4 NMs enhance the T cell-mediated anti-tumour immune response and DTX-CA4 NMs in combination with an immune checkpoint inhibitor, anti-PDL1 antibody, enhance the anti-tumour response. We additionally showed that DTX-CA4 NMs effectively attenuate the production of ceramide-1-phosphate, a key metabolite of the sphingolipid pathway, by downregulating the expression of ceramide kinase at both transcriptional and translational levels. Therefore, this study presents the engineering of effective DTX-CA4 NMs for targeting the tumour microenvironment that can be explored further for clinical applications.

Lithocholic acid derivatives as potent modulators of the nuclear receptor RORγt.

Retinoic acid receptor-related orphan receptor γt (RORγt) is a nuclear receptor found in various tissues that plays a crucial role in the differentiation and proliferation of T helper 17 (Th17) cells, as well as in their generation of the pro-inflammatory cytokine IL-17A. RORγt represents a promising therapeutic target for autoimmune diseases, metabolic disorders, and multiple tumors. Despite extensive research efforts focused on the development of small molecule RORγt modulators, no drug candidates have advanced to phase 3 clinical trials owing to a lack of efficacy or safety margin. This outcome highlights the unmet need to optimize small molecule drug candidates targeting RORγt to develop effective therapies for autoimmune and inflammatory diseases. In this study, we synthesized and evaluated 3-oxo-lithocholic acid amidates as a new class of RORγt modulators. Our evaluation entailed biophysical screening, cellular screening in different platforms, molecular docking, and in vitro pharmacokinetic profiling. The top compound from our study (3-oxo-lithocholic acid amidate, A2) binds to RORγt at an equilibrium dissociation constant (KD) of 16.5 ± 1.34 nM based on microscale thermophoresis (MST). Assessment of the efficacy of A2 in the cellular RORγt reporter luciferase assay revealed a half-maximal inhibitory concentration (IC50) value of 225 ± 10.4 nM. Unlike 3-oxo-lithocholic acid, A2 demonstrated the ability to reduce the IL-17A mRNA expression levels in EL4 cells with RORγt expression using quantitative reverse transcriptase PCR (RT-PCR). Validation of the desirable physicochemical properties and stability of A2 sets the stage for the preclinical evaluation of this new class of RORγt modulators in animal models of autoimmune diseases.

Cholic Acid-Derived Gemini Amphiphile Can Eradicate Interkingdom Polymicrobial Biofilms and Wound Infections.

Biofilm infections are mainly caused by Gram-positive bacteria (GPB) like Staphylococcus aureus, Gram-negative bacteria (GNB) like Pseudomonas aeruginosa, and fungi like Candida albicans. These infections are responsible for antimicrobial tolerance, and commensal interactions of these microbes pose a severe threat to chronic infections. Treatment therapies against biofilm infections are limited to eradicating only 20-30% of infections. Here, we present the synthesis of a series of bile acid-derived molecules using lithocholic acid, deoxycholic acid, and cholic acid where two bile acid molecules are tethered through 3'-hydroxyl or 24'-carboxyl terminals with varying spacer length (trimethylene, pentamethylene, octamethylene, and dodecamethylene). Our structure-activity relationship investigations revealed that G21, a cholic acid-derived gemini amphiphile having trimethylene spacer tethered through the C24 position, is a broad-spectrum antimicrobial agent. Biochemical studies witnessed that G21 interacts with negatively charged lipoteichoic acid, lipopolysaccharide, and phosphatidylcholine moieties of GPB, GNB, and fungi and disrupts the microbial cell membranes. We further demonstrated that G21 can eradicate polymicrobial biofilms and wound infections and prevent bacteria and fungi from developing drug resistance. Therefore, our findings revealed the potential of G21 as a versatile antimicrobial agent capable of effectively targeting polymicrobial biofilms and wound infections, suggesting that it is a promising antimicrobial agent for future applications.

(Invited) A Flexible Anti-Biofilm Hygiene Coating for Water Devices

Biofilm is a microbiome complex comprising different bacterial colonies that typically adhere to device surfaces in water, which causes serious medical issues such as indwelling infections and outbreaks. Here we introduce a non-nanoparticle, flexible anti-biofilm hygiene coating consisting of lithocholic acid (LCA), zinc pyrithione (Zn) and cinnamaldehyde (Cn) (named as LCA-Zn-Cn) that largely prevents the bacteria adhesion to various water device surfaces such as stainless steel and glass through a synergistic mechanism (ACS Applied Bio Materials, 5 (2022) 3991). The existing chelated groups on LCA and Cn attract plenty of bacteria via hydrophobic interaction. Both bactericidal reaction by grafting biocidal groups from both LCA and Cn and bacteriostatic reaction by inhibiting cell division via zinc ions (Zn) lead to a largely improved bacteria/biofilm prevention. The antibacterial performance was assessed by using JIS Z 2801/ISO 22196 method. The designed LCA-Zn-Cn coating displayed log10 reduction of 4.23 (99.9% reduction) of E. coli and log10reduction of 3.51 (99.8% reduction) of E. faecalis on stainless steel, which are much higher than the control samples, demonstrating a promising colonization inhibition. Due to the strong crosslinked configuration, the coating still showed >90.9% bacterial reduction after 3000 abrasion cycles and over 99.9% bacteria reduction after a high flow velocity of 1.99 m/s test, which confirmed the enhanced mechanical durability. By applying either spray or dip-coating, the designed polymer composite can be coated on a variety of irregular water devices with mass production using auto-controlled robot arm. The desired hybrid coating provides an effective solution to prevent bacterium and biofilm formation on flexible water devices. Keywords: Anti-biofilm; Anti-bacteria; Coating; Water devices; Flexibility

Ethnobotany and phytochemistry of plants used to treat musculoskeletal disorders among Skaw Karen, Thailand

Context Musculoskeletal system disorders (MSD) are prevalent around the world affecting the health of people, especially farmers who work hard in the field. Karen farmers use many medicinal plants to treat MSD. Objective This study collects traditional plant-based remedies used by the Skaw Karen to treat MSD and evaluates their active phytochemical compounds. Materials and methods The ethnobotanical study was conducted in six Karen villages in Chiang Mai province using semi-structured interviews were of 120 informants. The data were analyzed using ethnobotanical indices including use values (UV), choice value (CV), and informant consensus factor (ICF). Consequently, the 20 most important species, according to the indices, were selected for phytochemical analysis using LC–MS/MS. Results A total of 3731 use reports were obtained for 139 species used in MSD treatment. The most common ailments treated with those plants were muscular pain. A total of 172 high-potential active compounds for MSD treatment were identified. Most of them were flavonoids, terpenoids, alkaloids, and steroids. The prevalent phytochemical compounds related to treat MSD were 9-hydroxycalabaxanthone, dihydrovaltrate, morroniside, isoacteoside, lithocholic acid, pomiferin, cucurbitacin E, leonuriside A, liriodendrin, and physalin E. Sambucus javanica Reinw. ex Blume (Adoxaceae), Betula alnoides Buch.-Ham. ex D.Don (Betulaceae), Blumea balsamifera (L.) DC. (Asteraceae), Plantago major L. (Plantaginaceae) and Flacourtia jangomas (Lour.) Raeusch. (Salicaceae) all had high ethnobotanical index values and many active compounds. Discussion and conclusions This study provides valuable information, demonstrating low-cost medicine plants that are locally available. It is a choice of treatment for people living in remote areas.

Lithocholic acid decreases mRNA stability of nuclear receptor PPARα by upregulating miR-21 expression in hepatoma HepG2 cells

To investigate the regulatory effects of lithocholic acid (LCA) on nuclear receptor peroxisome proliferatoractivated receptor-alpha (PPARα) mRNA stability at the post-transcriptional level. PPARα 3'UTR luciferase reporter gene vectors were constructed and transfected into HepG2 cells to observe the changes in cellular luciferase activity in response to LCA treatments. Bioinformatic prediction and miRNA PCR array technique were used to identify the differentially expressed miRNAs induced by LCA and their potential binding sites on the 3'UTR. The binding sites (Mut1, Mut2 and Mut1+Mut2) were mutated to compare the changes in cellular luciferase activity following LCA treatment. Western blotting and RTqPCR were used to detect the activated signaling pathway and the expression levels of its downstream transcription factors in LCA-treated cells. The changes in PPARα protein expression level were detected in the cells following overexpression of the transcription factors. Treatment with 100 μmol/L LCA significantly reduced luciferase activity of PPARα 3'UTR1 and 3'UTR2 in HepG2 cells by more than 50% (P<0.01) and induced significant upregulation of miR-21 and miR-22, especially the former (by 2.35 folds, P<0.05). Two predicted miR-21-binding sites in the 3'UTR1 were mutated to construct Mut1, Mut2 and Mut1+Mut2 reporter gene vectors. LCA treatment down-regulated 3'UTR1 luciferase activity by 51%, while Mut1, Mut2, and Mut1+Mut2 were down-regulated by 37%, 39%, and 13%, respectively. LCA caused ERK1/2 phosphorylation and activation of the ERK1/2 signaling pathway, and treatment with 100 μmol/L LCA upregulated the expression of transcription factor early growth response 1 (EGR1) by 5.83 folds (P<0.01). Transient overexpression of EGR1 significantly decreased cellular PPARα protein levels (P<0.05). LCA reduces PPARα mRNA stability and thus decreases PPARα mRNA and protein expressions in hepatocytes by activating the ERK1/2 signaling pathway and upregulating EGR1 and miR-21, which targets 3'UTR regulatory region of PPARα mRNA.

Integrated rumen microbiome and serum metabolome analysis responses to feed type that contribution to meat quality in lambs

BackgroundLifestyle factors, such as diet, are known to be a driver on the meat quality, rumen microbiome and serum metabolites. Rumen microbiome metabolites may be important for host health, the correlation between rumen microbiome and production of rumen metabolites are reported, while the impact of rumen microbiome on the serum metabolome and fatty acid of meat are still unclear. This study was designed to explore the rumen microbiome, serum metabolome and fatty acid of meat in response to the grass diet and concentrate diet to lambs, and the relationship of which also investigated.MethodsIn the present study, 12 lambs were randomly divided into two groups: a grass diet (G) and a concentrate diet (C). Here, multiple physicochemical analyses combined with 16S rRNA gene sequences and metabolome analysis was performed to reveal the changes that in response to feed types.ResultsThe concentrate diet could improve the growth performance of lambs compared to that fed with the grass diet. The microbiome composition was highly individual, compared to the concentrate group, the abundance of Rikenellaceae_RC9_gut_group, F082_unclassified, Muribaculaceae_unclassified, Ruminococcaceae_NK4A214_group, Bacteroidetes_unclassified, and Bacteroidales_UCG-001_unclassified were significantly (P < 0.05) lower in the grass group, while, the abundance of Succinivibrio, Succinivibrionaceae_UCG-002, Fibrobacter and Christensenellaceae_R-7_group were significantly (P < 0.05) higher in the grass group. Serum metabolomics analysis combined with enrichment analysis revealed that serum metabolites were influenced by feed type as well as the metabolic pathway, and significantly affected serum metabolites involved in amino acids, peptides, and analogues, bile acids, alcohols and derivatives, linoleic acids derivatives, fatty acids and conjugates. Most of the amino acids, peptides, and analogues metabolites were positively associated with the fatty acid contents. Among the bile acids, alcohols and derivatives metabolites, glycocholic was positively associated with all fatty acid contents, except C18:0, while 25-Hydroxycholesterol and lithocholic acid metabolites were negatively associated with most of the fatty acid contents.ConclusionCorrelation analysis of the association of microbiome with metabolite features, metabolite features with fatty acid provides us with comprehensive understanding of the composition and function of microbial communities. Associations between utilization or production were widely identified among affected microbiome, metabolites and fatty acid, and these findings will contribute to the direction of future research in lamb.

Exploring the effect and mechanism of cucurbitacin B on cholestatic liver injury based on network pharmacology and experimental verification

Ethnopharmacological relevanceCholestatic liver injury (CLI) is a pathologic process with the impairment of liver and bile secretion and excretion, resulting in an excessive accumulation of bile acids within the liver, which leads to damage to both bile ducts and hepatocytes. This process is often accompanied by inflammation. Cucumis melo L is a folk traditional herb for the treatment of cholestasis. Cucurbitacin B (CuB), an important active ingredient in Cucumis melo L, has significant anti-inflamamatory effects and plays an important role in diseases such as neuroinflammation, skin inflammation, and chronic hepatitis. Though numerous studies have confirmed the significant therapeutic effect of CuB on liver diseases, the impact of CuB on CLI remains uncertain. Consequently, the objective of this investigation is to elucidate the therapeutic properties and potential molecular mechanisms underlying the effects of CuB on CLI. Aim of the studyThe aim of this paper was to investigate the potential protective mechanism of CuB against CLI. MethodsFirst, the corresponding targets of CuB were obtained through the SwissTargetPrediction and SuperPre online platforms. Second, the DisGeNET database, GeneCards database, and OMIM database were utilized to screen therapeutic targets for CLI. Then, protein-protein interaction (PPI) was determined using the STRING 11.5 data platform. Next, the OmicShare platform was employed for the purpose of visualizing the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. The molecular docking technique was then utilized to evaluate the binding affinity existing between potential targets and CuB. Subsequently, the impacts of CuB on the LO2 cell injury model induced by Lithocholic acid (LCA) and the CLI model induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) were determined by evaluating inflammation in both in vivo and in vitro settings. The potential molecular mechanism was explored by real-time quantitative polymerase chain reaction (RT-qPCR) and western blot (WB) techniques. ResultsA total of 122 CuB targets were collected and high affinity targets were identified through the PPI network, namely TLR4, STAT3, HIF1A, and NFKB1. GO and KEGG analyses indicated that the treatment of CLI with CuB chiefly involved the inflammatory pathway. In vitro study results showed that CuB alleviated LCA-induced LO2 cell damage. Meanwhile, CuB reduced elevated AST and ALT levels and the release of inflammatory factors in LO2 cells induced by LCA. In vivo study results showed that CuB could alleviate DDC-induced pathological changes in mouse liver, inhibit the activity of serum transaminase, and suppress the liver and systemic inflammatory reaction of mice. Mechanically, CuB downregulated the IL-6, STAT3, and HIF-1α expression and inhibited STAT3 phosphorylation. ConclusionBy combining network pharmacology with in vivo and in vitro experiments, the results of this study suggested that CuB prevented the inflammatory response by inhibiting the IL-6/STAT3/HIF-1α signaling pathway, thereby demonstrating potential protective and therapeutic effects on CLI. These results establish a scientific foundation for the exploration and utilization of natural medicines for CLI.

Modulation of In Vitro Macrophage Responses via Primary and Secondary Bile Acids in Dogs

Bile acids (BA) are important metabolites secreted into the intestinal lumen and impacted by luminal microbes and dietary intake. Prior studies in humans and rodents have shown that BAs are immunologically active and that primary and secondary BAs have distinct immune properties. Therefore, the composition of the gut BA pool may influence GI inflammatory responses. The current study investigated the relative immune modulatory properties of primary (cholic acid, CA) and secondary BAs (lithocholic acid, LCA) by assessing their effects on canine macrophage cytokine secretion and BA receptor (TGR5) expression. In addition, RNA sequencing was used to further interrogate how CA and LCA differentially modulated macrophage responses to LPS (lipopolysaccharide). We found that exposure to either CA or LCA influenced LPS-induced cytokine production via macrophages similarly, with suppression of TNF-α secretion and enhancement of IL-10 secretion. Neither BA altered the expression of the BA receptor TGR5. Transcriptomic analysis revealed that CA activated inflammatory signaling pathways in macrophages involving type II interferon signaling and the aryl hydrocarbon receptor, whereas LCA activated pathways related to nitric oxide signaling and cell cycle regulation. Thus, we concluded that both primary and secondary BAs are active modulators of macrophage responses in dogs, with differential and shared effects evident with sequencing analysis.