De novo production of protoberberine and benzophenanthridine alkaloids through metabolic engineering of yeast.

Protoberberine alkaloids and benzophenanthridine alkaloids (BZDAs) are subgroups of benzylisoquinoline alkaloids (BIAs), which represent a diverse class of plant-specialized natural metabolites with many pharmacological properties. Microbial biosynthesis has been allowed for accessibility and scalable production of high-value BIAs. Here, we engineer Saccharomyces cerevisiae to de novo produce a series of protoberberines and BZDAs, including palmatine, berberine, chelerythrine, sanguinarine and chelirubine. An ER compartmentalization strategy is developed to improve vacuole protein berberine bridge enzyme (BBE) activity, resulting in >200% increase on the production of the key intermediate (S)-scoulerine. Another promiscuous vacuole protein dihydrobenzophenanthridine oxidase (DBOX) has been identified to catalyze two-electron oxidation on various tetrahydroprotoberberines at N7-C8 position and dihydrobenzophenanthridine alkaloids. Furthermore, cytosolically expressed DBOX can alleviate the limitation on BBE. This study highlights the potential of microbial cell factories for the biosynthesis of a diverse group of BIAs through engineering of heterologous plant enzymes.

Biosynthetic Strategies of Berberine Bridge Enzyme-like Flavoprotein Oxidases toward Structural Diversification in Natural Product Biosynthesis.

Berberine bridge enzyme-like oxidases are often involved in natural product biosynthesis and are seen as essential enzymes for the generation of intricate pharmacophores. These oxidases have the ability to transfer a hydride atom to the FAD cofactor, which enables complex substrate modifications and rearrangements including (intramolecular) cyclizations, carbon-carbon bond formations, and nucleophilic additions. Despite the diverse range of activities, the mechanistic details of these reactions often remain incompletely understood. In this Review, we delve into the complexity that BBE-like oxidases from bacteria, fungal, and plant origins exhibit by providing an overview of the shared catalytic features and emphasizing the different reactivities. We propose four generalized modes of action by which BBE-like oxidases enable the synthesis of natural products, ranging from the classic alcohol oxidation reactions to less common amine and amide oxidation reactions. Exploring the mechanisms utilized by nature to produce its vast array of natural products is a subject of considerable interest and can lead to the discovery of unique biochemical activities.

Effect of the presence of berberine/curcumin on the binding of limonin to human serum albumin and antitumor activity in vitro.

The competition among drugs for binding to plasma proteins is regarded as a pharmacokinetic drug interaction. Competition between antitumor agents and other drugs for plasma protein binding can alter the free concentration of the drug, potentially impacting its efficacy and increasing the risk of toxic side effects. Through a range of spectroscopic techniques, this study examined the interaction between limonin and human serum albumin (HSA) in the context of berberine (Ber) and curcumin (Cur) under physiological conditions to clarify the binding mechanisms of binary and ternary systems at the molecular level. As demonstrated by fluorescence quenching experiments, Static quenching was identified as the mechanism of interaction between HSA and limonin. The results of site competition experiments indicated that the binding site between limonin and HSA was site I, a result further supported by molecular docking simulations. Through the use of thermodynamic data calculations, it was determined that limonin forms a stable complex with HSA by establishing hydrogen bonds and van der Waals forces. Circular dichroism (CD) spectroscopy, three-dimensional (3D) fluorescence spectroscopy, and synchronous fluorescence spectroscopy (SFS) employed to validate the notion that limonin perturbed the microenvironment of amino acids and induced conformational changes in HSA. What's more, the presence of Ber or Cur was found to have further modified the alterations observed in the interaction between the original HSA-limonin binary system. In vitro cellular experiments showed that interaction with HSA reduced the antitumor activity of limonin. In contrast, adding Ber or Cur increased the inhibition rate of tumor cells. The coexistence of both Ber and Cur significantly diminished limonin's binding affinity to HSA. The current investigation enhances comprehension regarding the binding characteristics and interaction mechanisms involving limonin, Ber, Cur, and HSA. It explores the potential of HSA as a versatile drug carrier and furnishes theoretical underpinnings for co-administrative strategies.

Isolation of Protoplasts from Tissues of Mexican Prickly Poppy (Argemone mexicana L.): An Alkaloid-Producing Medicinal Plant.

Protoplasts are plant cells from which the pectocellulosic cell wall has been removed, thus keeping the plasma membrane intact. For plant secondary metabolites research, this system is a powerful tool to study the metabolites' dynamics inside the cells, such as the subcellular localization of proteins, characterization of gene function, transcription factors involved in metabolite pathways, protein transport machinery, and to perform single-cell omics studies. Due to its lack of a cell wall, better images of the interior of the cell can be obtained compared to the whole tissue. This allows the identification of specific cell types involved in the accumulation of specialized metabolites, such as alkaloids, given their autofluorescence properties. Here is a simplified protocol to obtain protoplasts from leaves and in vitro cell cultures from Argemone mexicana, which produces the pharmacologically important alkaloids berberine and sanguinarine.

Berberine analog of chloramphenicol exhibits a distinct mode of action and unveils ribosome plasticity.

Chloramphenicol (CHL) is an antibiotic targeting the peptidyl transferase center in bacterial ribosomes. We synthesized a new analog, CAM-BER, by substituting the dichloroacetyl moiety of CHL with a positively charged aromatic berberine group. CAM-BER suppresses bacterial cell growth, inhibits protein synthesis in vitro, and binds tightly to the 70S ribosome. Crystal structure analysis reveals that the bulky berberine group folds into the P site of the peptidyl transferase center (PTC), where it competes with the formyl-methionine residue of the initiator tRNA. Our toe-printing data confirm that CAM-BER acts as a translation initiation inhibitor in stark contrast to CHL, a translation elongation inhibitor. Moreover, CAM-BER induces a distinct rearrangement of conformationally restrained nucleotide A2059, suggesting that the 23S rRNA plasticity is significantly higher than previously thought. CAM-BER shows potential in avoiding CHL resistance and presents opportunities for developing novel berberine derivatives of CHL through medicinal chemistry exploration.

Interactions of three berberine mid-chain fatty acid salts with bovine serum albumin (BSA): Spectroscopic analysis and molecular docking.

In this paper, the interaction of three berberine mid-chain fatty acid salts ([BBR][FAs]), viz. berberine caproate ([BBR][CAP]), berberine heptylate ([BBR][HEP]) and berberine octoate ([BBR][OCT]), with bovine serum albumin (BSA) was studied by means of UV-visible absorption spectroscopy, fluorescence spectroscopy, fourier transform infrared spectroscopy (FT-IR) and molecular docking techniques. Fluorescence experiments revealed that three berberine salts quench the fluorescence of BSA by static quenching mechanism resulted from a stable [BBR][FAs]-BSA complex formation. The stoichiometric numbers of [BBR][FAs]-BSA complexes were found to be 1:1. Synchronous and three-dimensional fluorescence spectra as well as FT-IR demonstrated that the binding of [BBR][FAs] altered the microenvironment and conformation of BSA. The binding average distance from [BBR][FAs] to BSA (3.2-3.5 nm) was determined according to Förster energy transfer theory. Site probe investigation showed that [BBR][FAs] bound to BSA active site I (sub-domain IIA). The binding promotes the esterase-like activity of BSA. The molecular docking results confirmed the fluorescence competition findings and provided the type of binding forces. Furthermore, the relationship between the anionic chain length of [BBR][FAs] and the interaction was explored, and the positive correlation was found.

Differences in Metabolite Profiles of Dihydroberberine and Micellar Berberine in Caco-2 Cells and Humans-A Pilot Study.

We investigated the pharmacokinetic pathway of berberine and its metabolites in vitro, in Caco-2 cells, and in human participants following the administration of dihydroberberine (DHB) and micellar berberine (LipoMicel®, LMB) formulations. A pilot trial involving nine healthy volunteers was conducted over a 24 h period; blood samples were collected and subjected to Ultra High-Performance Liquid Chromatography-High Resolution Mass Spectrometry (UHPLC-HRMS) analyses to quantify the concentrations of berberine and its metabolites. Pharmacokinetic correlations indicated that berberrubine and thalifendine follow distinct metabolic pathways. Additionally, jatrorrhizine sulfate appeared to undergo metabolism differently compared to the other sulfated metabolites. Moreover, berberrubine glucuronide likely has a unique metabolic pathway distinct from other glucuronides. The human trial revealed significantly higher blood concentrations of berberine metabolites in participants of the DHB treatment group compared to the LMB treatment group-except for berberrubine glucuronide, which was only detected in the LMB treatment group. Similarly, results from in vitro investigations showed significant differences in berberine metabolite profiles between DHB and LMB. Dihydroberberine, dihydroxy-berberrubine/thalifendine and jatrorrhizine sulfate were detected in LMB-treated cells, but not in DHB-treated cells; thalifendine and jatrorrhizine-glucuronide were detected in DHB-treated cells only. While DHB treatment provided higher blood concentrations of berberine and most berberine metabolites, both in vitro (Caco-2 cells) and in vivo human studies showed that treatment with LMB resulted in a higher proportion of unmetabolized berberine compared to DHB. These findings suggest potential clinical implications that merit further investigation in future large-scale trials.

Natural compound library screening to identify berberine as a treatment for rheumatoid arthritis.

OBJECTIVE: Fibroblast-like synoviocytes (FLS) play a critical role on the exacerbation and deterioration of rheumatoid arthritis (RA). Aberrant activation of FLS pyroptosis signaling is responsible for the hyperplasia of synovium and destruction of cartilage of RA. This study investigated the screened traditional Chinese medicine berberine (BBR), an active alkaloid extracted from the Coptis chinensis plant, that regulates the pyroptosis of FLS and secretion of inflammatory factors in rheumatoid arthritis. METHODS: First, BBR was screened using a high-throughput drug screening strategy, and its inhibitory effect on RA-FLS was verified by in vivo and in vitro experiments. Second, BBR was intraperitoneally administrated into the collagen-induced arthritis rat model, and the clinical scores, arthritis index, and joint HE staining were evaluated. Third, synovial tissues of CIA mice were collected, and the expression of NLRP3, cleaved-caspase-1, GSDMD-N, Mst1, and YAP was detected by Western blot. RESULTS: The administration of BBR dramatically alleviated the severity of collagen-induced arthritis rat model with a decreased clinical score and inflammation reduction. In addition, BBR intervention significantly attenuates several pro-inflammatory cytokines (interleukin-1ß, interleukin-6, interleukin-17, and interleukin-18). Moreover, BBR can reduce the pyroptosis response (caspase-1, NLR family pyrin domain containing 3, and gasdermin D) of the RA-FLS in vitro, activating the Hippo signaling pathway (Mammalian sterile 20-like kinase 1, yes-associated protein, and transcriptional enhanced associate domains) so as to inhibit the pro-inflammatory effect of RA-FLS. CONCLUSION: These results support the role of BBR in RA and may have therapeutic implications by directly repressing the activation, migration of RA-FLS, which contributing to the attenuation of the progress of CIA. Therefore, targeting PU.1 might be a potential therapeutic approach for RA. Besides, BBR inhibited RA-FLS pyroptosis by downregulating of NLRP3 inflammasomes (NLRP3, caspase-1) and eased the pro-inflammatory activities via activating the Hippo signaling pathway, thereby improving the symptom of CIA.

Berberine attenuates cognitive dysfunction and hippocampal apoptosis in rats with prediabetes.

The cognitive dysfunction caused by prediabetes causes great difficulties in human life, and the terrible thing is that the means to prevent the occurrence of this disease are very limited at present, Berberine has shown the potential to treat diabetes and cognitive dysfunction, but it still needs to be further explored to clarify the mechanism of its therapeutic effect. Therefore, the aim of this study was to investigate the effects and mechanisms of Berberine on prediabetes-induced cognitive dysfunction. Prediabetes rat model was induced by a high-fat diet and a normal diet was used as a control. They were fed for 20 weeks. At week 13, the model rats were given 100 mg/kg Berberine by gavage for 7 weeks. The cognitive function of rats was observed. At the same time, OGTT, fasting blood glucose, blood lipids, insulin and other metabolic parameters, oxidative stress, and apoptosis levels were measured. The results showed that the model rats showed obvious glucose intolerance, elevated blood lipids, and insulin resistance, and the levels of oxidative stress and apoptosis were significantly increased. However, after the administration of Berberine, the blood glucose and lipid metabolism of prediabetic rats were significantly improved, and the oxidative stress level and apoptosis level of hippocampal tissue were significantly reduced. In conclusion, Berberine can alleviate the further development of diabetes in prediabetic rats, reduce oxidative stress and apoptosis in hippocampal tissue, and improve cognitive impairment in prediabetic rats.

Cationic fructan-based pH and intestinal flora dual stimulation nanoparticle with berberine for targeted therapy of IBD.

Inflammatory bowel disease (IBD) can cause intestinal microbial imbalance and aggravate intestinal inflammation. Mixed fructan is more easily fermented by colonic microorganisms and can be used as colonic drug delivery materials. Here, we constructed a mixed fructan based nanoparticle with dual targeted stimulation of pH and intestinal flora to effectively deliver berberine for the treatment of ulcerative colitis (UC). The complex of fructan based nanoparticle and berberine (BBRNPs) significantly ameliorated the inflammatory response of sodium dextran sulfate (DSS)-induced colitis in mice by inhibiting the activation of NF-κB/STAT-3 pathway and increasing tight junction protein expression in vivo. Importantly, BBRNPs improved the responsiveness of colitis microbiome and effectively regulated the relative homeostasis of harmful flora Enterobacteriaceae and Escherichia-shigolla, and beneficial flora Ruminococcaceae and Akkermansiaceae. This study provides a promising strategy for the effective treatment of UC and expands the application of branched fructan in pharmaceutics.

Integrated non-targeted metabolomics and network pharmacology to reveal the mechanisms of berberine in the long-term treatment of PTZ-induced epilepsy.

AIMS: The increasing resistance to anti-seizure medications (ASMs) and the ambiguous mechanisms of epilepsy highlight the pressing demand for the discovery of pioneering lead compounds. Berberine (BBR) has received significant attention in recent years within the field of chronic metabolic disorders. However, the reports on the treatment of epilepsy with BBR are not systematic and the mechanism remains unclear. MAIN METHODS: In this study, the seizure behaviors of mice were recorded following subcutaneous injection of pentetrazol (PTZ). Non-targeted metabolomics was used to analyze the serum metabolites based on ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). Meanwhile, multivariate statistical methods were used for metabolite identification and pathway analysis. Furthermore, network pharmacology, molecular docking, and quantitative real-time PCR assay were used for the target identification. KEY FINDINGS: BBR had anti-seizure effects on PTZ-induced seizure mice after long-term treatment. Tryptophan metabolism and phenylalanine metabolism were involved in regulating the therapeutic effects of BBR. SIGNIFICANCE: This study reveals the potential mechanism of BBR for epilepsy treatment based on non-targeted metabolomics and network pharmacology, which provides evidence for uncovering the pathogenesis of epilepsy, suggesting that BBR is a potential lead compound for anti-epileptic treatment.

Berberine exerts antidepressant effects in vivo and in vitro through the PI3K/AKT/CREB/BDNF signaling pathway.

BACKGROUND: Depression, a global neuropsychiatric disorder, brings a serious burden to patients and society as its incidence continues to rise. Berberine is one of the main compounds of a variety of Chinese herbal medicines and has been shown to have multiple pharmacological effects. However, whether berberine can exert antidepressant effects in vivo and in vitro and its related mechanisms remain to be explored. METHODS: The chronic restraint stress (CRS) method and corticosterone (CORT) were applied to simulate depression-like behavior in vivo and neuronal apoptosis in vitro, respectively. The antidepressant effects of berberine were evaluated by behavioral tests and changes in the content of monoamine neurotransmitters. Inflammatory cytokines were detected and immunofluorescence staining was used to observe the expression levels of apoptosis-related proteins. RT-qPCR and Western blot were used to examine the mRNA and protein expression (or phosphorylation) levels of biomarkers of the PI3K/AKT/CREB/BDNF signaling pathways. RESULTS: Behavioral tests and levels of neurotransmitters proved that berberine could effectively ameliorate depression-like symptoms in CRS mice. Meanwhile, the results of ELISA and immunofluorescence staining showed that berberine could alleviate inflammatory status and reduce cell apoptosis in vivo and in vitro. Moreover, the changes of the PI3K/AKT/CREB/BDNF signaling pathway induced by CRS or CORT in mouse hippocampus or HT-22 cells were significantly reversed by berberine. CONCLUSION: Our current study suggested that berberine could exert antidepressant effects in vitro and in vivo, which may be associated with the PI3K/AKT/CREB/BDNF signaling pathway.

Research progress of natural active compounds on improving podocyte function to reduce proteinuria in diabetic kidney disease.

Diabetic kidney disease (DKD) is a primary cause of end-stage renal disease. Proteinuria is a clinical indicator of the different stages of DKD, and podocyte injury is a major cause of proteinuria. Podocyte-specific proteins (PSPs) play important roles in the normal filtration of podocytes. Studies have shown that natural active compounds (NACs) can ameliorate proteinuria; however, the mechanism related to PSPs needs to be explored. In this study, the five stages of DKD related to proteinuria and the functions of PSPs are displayed separately. Mechanisms for ameliorating proteinuria and improving the PSPs of the 15 NACs are summarized. The in vitro and in vivo mechanistic research showed that five compounds, astragaloside IV, ligustrazine, berberine, emodin and resveratrol, exerted renal protective effects via AMPK signaling, icariin and berberine via TLR4 signaling, hirudin and baicalin via MAPK signaling, curcumin and baicalin via NF-κB signaling, and emodin via protein kinase RNA-like endoplasmic reticulum kinase signaling. The 13 PSPs were divided into five categories: actin cytoskeleton, basal domain, apical domain, slit diaphragm, and others. In conclusion, anti-inflammatory effects, anti-oxidative stress, and enhanced autophagy are the main mechanisms underlying the ameliorative effects of NACs. Podocyte apoptosis is mainly related to nephrin and podocin, which are the most studied slit diaphragm PSPs.

Berberine Protects against High-Energy and Low-Protein Diet-Induced Hepatic Steatosis: Modulation of Gut Microbiota and Bile Acid Metabolism in Laying Hens.

Berberine (BBR) is a natural alkaloid with multiple biotical effects that has potential as a treatment for fatty liver hemorrhagic syndrome (FLHS). However, the mechanism underlying the protective effect of BBR against FLHS remains unclear. The present study aimed to investigate the effect of BBR on FLHS induced by a high-energy, low-protein (HELP) diet and explore the involvement of the gut microbiota and bile acid metabolism in the protective effects. A total of 90 healthy 140-day-old Hy-line laying hens were randomly divided into three groups, including a control group (fed a basic diet), a HELP group (fed a HELP diet), and a HELP+BBR group (high-energy, high-protein diet supplemented with BBR instead of maize). Our results show that BBR supplementation alleviated liver injury and hepatic steatosis in laying hens. Moreover, BBR supplementation could significantly regulate the gut's microbial composition, increasing the abundance of Actinobacteria and Romboutsia. In addition, the BBR supplement altered the profile of bile acid. Furthermore, the gut microbiota participates in bile acid metabolism, especially taurochenodeoxycholic acid and α-muricholic acid. BBR supplementation could regulate the expression of genes and proteins related to glucose metabolism, lipid synthesis (FAS, SREBP-1c), and bile acid synthesis (FXR, CYP27a1). Collectively, our findings demonstrate that BBR might be a potential feed additive for preventing FLHS by regulating the gut microbiota and bile acid metabolism.

DFT, ADME studies and evaluation of the binding with HSA and MAO-B inhibitory potential of protoberberine alkaloids from Guatteria friesiana: theoretical insights of promising candidates for the treatment of Parkinson's disease.

CONTEXT: Parkinson's disease is a chronic neurodegenerative condition that has no cure, characterized by the progressive degeneration of specific brain cells responsible for producing dopamine, a crucial neurotransmitter for controlling movement and muscle coordination. Parkinson's disease is estimated to affect around 1% of the world's population over the age of 60, but it can be diagnosed at younger ages. One of the treatment strategies for Parkinson's disease involves the use of drugs that aim to increase dopamine levels or simulate the action of dopamine in the brain. A class of commonly prescribed drugs are the so-called monoamine oxidase B (MAO-B) inhibitors due to the fact that this enzyme is responsible for metabolizing dopamine, thus reducing its levels in the brain. Studies have shown that berberine-derived alkaloids have the ability to selectively inhibit MAO-B activity, resulting in increased dopamine availability in the brain. In this context, berberine derivatives 13-hydroxy-discretinine and 7,8-dihydro-8-hydroxypalmatine, isolated from Guatteria friesiana, were evaluated via density functional theory followed by ADME studies, docking and molecular dynamic simulations with MAO-B, aiming to evaluate their anti-Parkinson potential, which have not been reported yet. Docking simulations with HSA were carried out aiming to evaluate the transport of these molecules through the circulatory system. METHODS: The 3D structures of the berberine-derived alkaloids were modeled via the DFT approach at B3LYP-D3(BJ)/6-311 + + G(2df, 2pd) theory level using Gaussian 09 software. Solvation free energies were determined through Truhlar's solvation model. MEP and ALIE maps were generated with Multiwfn software. Autodock Vina software was used for molecular docking simulations and analysis of the interactions in the binding sites. The 3D structure of MAO-B was obtained from the Protein Data Bank website under PDB code 2V5Z. For the interaction of studied alkaloids with human serum albumin (HSA) drug sites, 3D structures with PDB codes 2BXD, 2BXG, and 4L9K were used. Molecular dynamics simulations were carried out using GROMACS 2019.4 software, with the GROMOS 53A6 force field at 100 ns simulation time. The estimation of the ligand's binding free energies was obtained via molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) method.

Binding of berberine to PEBP1 synergizes with sorafenib to induce the ferroptosis of hepatic stellate cells.

Hepatic stellate cell (HSC) activation is the key process in hepatic fibrosis (HF) development. Targeted death of HSCs could be effective in the prevention and treatment of HF. Phosphatidylethanolamine-binding protein (PEBP)1 can trigger ferroptosis by mediating peroxide production, but how it modulates HSC ferroptosis is not known. We screened natural small molecules that could bind with PEBP1, and investigated the mechanism by which it promotes HSC ferroptosis. The maximum binding energy of berberine with PEBP1 was - 8.51 kcal/mol, indicating that berberine could bind strongly with PEBP1. Berberine binding to PEBP1 could promote HSC ferroptosis via synergy of its actions with those of sorafenib, but it could not induce ferroptosis alone. Combined administration of berberine enhanced the ferroptotic effects of low-dose sorafenib upon HSCs. Herein, we revealed that PEBP1 might be a target that could enhance the effects of sorafenib, which could provide a new therapeutic approach for HF treatment.

Berberine ameliorates renal interstitial inflammation and fibrosis in mice with unilateral ureteral obstruction.

Berberine acts via multiple pathways to alleviate fibrosis in various tissues and shows renoprotective effects. However, its role and underlying mechanisms in renal fibrosis remain unclear. Herein, we aimed to investigate the protective effects and molecular mechanisms of berberine against unilateral ureteric obstruction-induced renal fibrosis. The results indicated that berberine treatment (50 mg/kg/day) markedly alleviated histopathological alterations, collagen deposition and inflammatory cell infiltration in kidney tissue and restored mouse renal function. Mechanistically, berberine intervention inhibited NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome activation and the levels of the inflammatory cytokine IL-1ß in the kidneys of unilateral ureteric obstruction mice. In addition, berberine relieved unilateral ureteric obstruction-induced renal injury by activating adenosine monophosphate-activated protein kinase (AMPK) signalling and promoting fatty acid ß-oxidation. In vitro models showed that berberine treatment prevented the TGF-ß1-induced profibrotic phenotype of hexokinase 2 (HK-2) cells, characterized by loss of an epithelial phenotype (alpha smooth muscle actin [α-SMA]) and acquisition of mesenchymal marker expression (E-cadherin), by restoring abnormal fatty acid ß-oxidation and upregulating the expression of the fatty acid ß-oxidation related-key enzymes or regulators (phosphorylated-AMPK, peroxisome proliferator activated receptor alpha [PPARα] and carnitine palmitoyltransferase 1A [CPT1A]). Collectively, berberine alleviated renal fibrosis by inhibiting NLRP3 inflammasome activation and protected tubular epithelial cells by reversing defective fatty acid ß-oxidation. Our findings might be exploited clinically to provide a potential novel therapeutic strategy for renal fibrosis.

Berberine-loaded MSC-derived sEVs encapsulated in injectable GelMA hydrogel for spinal cord injury repair.

After spinal cord injury (SCI), local inflammatory response and fibrous scar formation severely hinder nerve regeneration. Berberine (Ber) has a powerful regulatory effect on the local microenvironment, but its limited solubility and permeability through the blood-brain barrier severely limit its systemic efficacy. Human umbilical cord mesenchymal stem cells (hUC-MSCs)-derived small extracellular vesicles (sEVs) are natural nanocarriers with high cargo loading capacity, and can cross the blood-brain barrier. Most importantly, sEVs can improve drug solubility and drug utilization. Therefore, they can overcome many defects of Ber application. This experiment aimed to design a Ber-carrying hUC-MSCs-derived sEVs and GelMA hydrogel. Ber was loaded into sEVs (sEVs-Ber) by ultrasonic co-incubation with a drug loading capacity (LC) of 15.07%. The unhindered release of up to 80% of sEVs-Ber from GelMA hydrogel was accomplished for up to 14 days. And they could be directly absorbed by local cells of injury, allowing for direct local delivery of the drug and enhancing its efficacy. The experimental results confirmed injecting GelMA-sEVs-Ber into spinal cord defects could exert anti-inflammatory effects by regulating the expression of inflammatory factors. It also demonstrated the anti-fibrotic effect of Ber in SCI for the first time. The modulatory effects of sEVs and Ber on the local microenvironment significantly promoted nerve regeneration and recovery of motor function in post-SCI rats. These results demonstrated that the GelMA-sEVs-Ber dual carrier system is a promising therapeutic strategy for SCI repair.

Comparing the Influences of Metformin and Berberine on the Intestinal Microbiota of Rats With Nonalcoholic Steatohepatitis.

BACKGROUND/AIM: High-fat diets induce shifts in the gut microbial community structure in patients or animals with non-alcoholic steatohepatitis (NASH). The objective of this study was to investigate the influence of metformin (MET) and berberine (BER) on the intestinal microbiota of rats with NASH. MATERIALS AND METHODS: Forty specific pathogen-free male Sprague-Dawley rats were randomized into 4 groups. Model rats were fed high-fat diets to create NASH models. MET or BER rats were administrated MET or BER, respectively, at the onset of induction of NASH. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), cholesterol, and triglycerides were examined. Plasma endotoxin levels were measured using the turbidimetric endotoxin assay. The incidence of bacterial translocation describes the passage of bacteria of the gastrointestinal tract through the intestinal mucosa barrier to mesenteric lymph nodes and other organs. Hematoxylin and eosin and oil red O staining were used for histopathological analysis. High throughput 16S rRNA sequencing was carried out for analyzing the composition of intestinal microbiota. RESULTS: High-fat diets caused NASH after 16-week induction. Administration of MET and BER ameliorated NASH by attenuating hepatic steatosis and inflammation and decreasing the plasma levels of endotoxin. MET and BER restored the composition of the intestinal microbiota disrupted by NASH. Both MET and BER altered the abundance of Atopobiaceae, Brevibacterium, Christensenellaceae, Coriobacteriales, Papillibacter, Pygmaiobacter, and Rikenellaceae RC9 in rats with NASH. The screened intestinal microbiota may be responsible for the improvement in fat accumulation and glucose metabolism. CONCLUSION: MET and BER demonstrated beneficial effects on the intestinal microbiota, which was disturbed in NASH. This finding may explain the functional mechanism of MET and BER in NASH.

Virtual screening and antimicrobial evaluation for identification of natural compounds as the prospective inhibitors of antibacterial drug resistance targets in Staphylococcus aureus.

Infectious diseases have remained a burgeoning cause of death and disability since long. Staphylococcus aureus (S. aureus) is a severe bacterial pathogen causing nosocomial and community infections. It exhibits widespread resistance to antibiotics posing a significant threat to their efficacy. For combating this challenge, different strategies may include modifying existing antibiotics, developing new antibacterial agents, and combining treatments with resistance mechanism inhibitors. Resistance in S. aureus occurs through horizontal gene transfer or chromosomal mutations. Acquisition mechanisms involve enzymatic modification, efflux, target bypass, and drug displacement. Mutations can impact drug targets, activate efflux pumps, or alter cell wall composition to impede drug access. Overcoming S. aureus resistance requires innovative approaches to preserve antibiotic effectiveness. The present study involves the virtual screening of phytochemicals of diverse chemical classes from Zinc database against the antibiotic resistant targets of S. aureus like ß-Lactamase, Penicillin Binding Protein 2a (PBP2a), Dihydrofolate reductase (DHFR), DNA gyrase, Multidrug ABC transporter SAV1866, Undecaprenyl diphosphate synthase (UPPS), etc. Thymol, eugenol, gallic acid, l-ascorbic acid, curcumin, berberine and quercetin were identified as potential molecules based on their docking score, binding interactions. These molecules were further analyzed for the ADMET and drug likeness properties using pkCSM, SwissADME and Qikprop tools. Further in vitro evaluation of these molecules against antibiotic-resistant strains of S. aureus, both alone and in combination with antibiotics revealed significant findings. Curcumin demonstrated the lowest MIC values (31.25-62.5 µg/ml) when tested individually. Thymol, berberine, and quercetin displayed MIC values within the range of 125-250 µg/ml, while eugenol and gallic acid exhibited MIC values ranging from 500 to 1000 µg/ml. Notably, thymol exhibited potent synergy with all four antibiotics against clinical isolates of S. aureus, with Fractional inhibitory concentration index (FICI) values consistently below 0.5, highlighting its exceptional antibacterial activity, especially in combination with amoxicillin.