Β-glucuronidase Activity Research Articles

Chang-wei-qing, a Chinese herbal formula, ameliorates colitis-associated tumour development via inhibiting NF-κB and STAT3 signalling pathway

Context: Chang-wei-qing (CWQ) is a Chinese herbal recipe with clinical efficacy. However, the molecular mechanism underlying its recognized therapeutic benefits against colorectal cancer is still elusive.Objective: To investigate the potential beneficial effects of CWQ in drug-induced colitis-associated cancer (CAC) model and its mechanistic involvements in this disease.Materials and methods: Colitis-associated cancer model was induced by azoxymethane (AOM) and dextran sulphate sodium (DSS). CWQ was administrated by gavage. Colon length and tumour size were determined after resection. The colitis was systematically scored. The microbiota and population of Faecalibacterium prausnitzii (F. prausnitzii) Hauduroy & Duncan was analysed by quantitative polymerase chain reaction (PCR). β-Glucuronidase, d-lactose and endotoxin were determined with commercially available kits. Pro-inflammatory cytokines were analysed in the colon tissues. Relative protein expressions were determined by Western blotting.Results: High concentration CWQ significantly restored the colon length, decreased tumour number and size (1.7 ± 0.6 vs. 2.8 ± 0.4 mm, p < 0.01) and reduced colitis score (11.8 ± 2.1 vs. 18.2 ± 2.3, p < 0.01). CWQ also suppressed expansion of F. prausnitzii population (0.029 ± 0.015% vs. 0.052 ± 0.019%, p < 0.01). CWQ greatly inhibited the activity of β-glucuronidase and leakage of d-lactose and endotoxin. Meanwhile, the pro-inflammatory cytokines were remarkably decreased in CAC mice in response to CWQ treatment. We further demonstrated that CWQ inhibited both NF-κB and STAT3 signalling.Conclusions: We for the first time demonstrated the antitumour properties of CWQ in vivo via inhibiting NF-κB and STAT3 signalling.

A Probe-Enabled Approach for the Selective Isolation and Characterization of Functionally Active Subpopulations in the Gut Microbiome

Commensal microorganisms in the mammalian gut play important roles in host health and physiology, but a central challenge remains in achieving a detailed mechanistic understanding of specific microbial contributions to host biochemistry. New function-based approaches are needed that analyze gut microbial function at the molecular level by coupling detection and measurements of in situ biochemical activity with identification of the responsible microbes and enzymes. We developed a platform employing β-glucuronidase selective activity-based probes to detect, isolate, and identify microbial subpopulations in the gut responsible for this xenobiotic metabolism. We find that metabolic activity of gut microbiota can be plastic and that between individuals and during perturbation, phylogenetically disparate populations can provide β-glucuronidase activity. Our work links biochemical activity with molecular-scale resolution without relying on genomic inference.

Lysosomal and network alterations in human mucopolysaccharidosis type VII iPSC-derived neurons

Mucopolysaccharidosis type VII (MPS VII) is a lysosomal storage disease caused by deficient β-glucuronidase (β-gluc) activity. Significantly reduced β-gluc activity leads to accumulation of glycosaminoglycans (GAGs) in many tissues, including the brain. Numerous combinations of mutations in GUSB (the gene that codes for β-gluc) cause a range of neurological features that make disease prognosis and treatment challenging. Currently, there is little understanding of the molecular basis for MPS VII brain anomalies. To identify a neuronal phenotype that could be used to complement genetic analyses, we generated two iPSC clones derived from skin fibroblasts of an MPS VII patient. We found that MPS VII neurons exhibited reduced β-gluc activity and showed previously established disease-associated phenotypes, including GAGs accumulation, expanded endocytic compartments, accumulation of lipofuscin granules, more autophagosomes, and altered lysosome function. Addition of recombinant β-gluc to MPS VII neurons, which mimics enzyme replacement therapy, restored disease-associated phenotypes to levels similar to the healthy control. MPS VII neural cells cultured as 3D neurospheroids showed upregulated GFAP gene expression, which was associated with astrocyte reactivity, and downregulation of GABAergic neuron markers. Spontaneous calcium imaging analysis of MPS VII neurospheroids showed reduced neuronal activity and altered network connectivity in patient-derived neurospheroids compared to a healthy control. These results demonstrate the interplay between reduced β-gluc activity, GAG accumulation and alterations in neuronal activity, and provide a human experimental model for elucidating the bases of MPS VII-associated cognitive defects.

2-Oxo-1,2,3,4-tetrahydropyrimidines Ethyl Esters as Potent β- Glucuronidase Inhibitors: One-pot Synthesis, In vitro and In silico Studies.

Glucuronidation is essential for the metabolism and excretion of toxic substances. β-Glucuronidase enzyme slows down the process of glucuronidation, and thus plays an important role in the on-set of colorectal carcinoma, and many other diseases. Inhibition of β- glucuronidase activity is thus identified as an important approach for the treatment of several diseases. Current study was aimed to synthesize a library of 2-oxo-1,2,3,4-tetrahydropyrimidine and to evaluate their β-glucuronidase inhibitory activity, and their mode of enzyme inhibition. We synthesized a series of 2-oxo-1,2,3,4-tetrahydropyrimidines 1-25 by fusing urea, ethyl acetoacetate, and a variety of aldehydes using copper nitrate trihydrate as catalyst. All synthesized compounds were evaluated for their in vitro β-glucuronidase inhibitory activity. In addition, molecular docking studies were also performed by using MOE docking tools. Eighteen compounds showed inhibitory activity better than the standard D-saccharic acid 1,4-lactone, a well known β-glucuronidase inhibitor (IC50 = 45.75 ± 2.16 µM). Compound 20 (IC50 = 1.36 ± 0.03 µM) showed an excellent inhibitory activity, thirty-five folds superior to the standard. Docking results highlighted the role of various chemical moieties at different positions on 2- oxo-1,2,3,4-tetrahydropyrimidine skeleton in enzyme inhibitory activity. This study has identified a class of potent β-glucuronidase inhibitors with the potential to be investigated further.

β-Glucuronidase and Its Relationship With Clinical Parameters and Biomarkers of Pesticide Exposure.

This study determined β-glucuronidase activity and its relationship with clinical, biochemical, and pesticide-exposure biomarkers in workers occupationally exposed mainly to OP insecticides. A cross-sectional study was conducted in pesticide applicators. Three study groups were established: reference group, moderate-exposure group, and high-exposure group. Blood samples were obtained to determine the activity of β-glucuronidase. Clinical analysis was conducted in a certified laboratory. The activity of β-glucuronidase was associated with clinical variables such as glucose, cholesterol, triglycerides, LDL, VLDL, HDL, atherogenic index, transaminases, and BuChE activity, but not with AChE activity. The activity of this enzyme exhibited a significant increase in the group of moderate exposure to pesticides but not in the group of high exposure. β-Glucuronidase activity is a sensitive biomarker under certain scenarios of OP exposure.

Serum β-glucuronidase activity as a biomarker for acute cholinesterase inhibitor pesticide poisoning

β-glucuronidase (BG) activity is a promising biomarker for diagnosis and prognosis after exposure to organophosphorous (OP) pesticides. The aim of this study was to evaluate the changes in serum BG activity in patients with acute OP poisoning and to determine whether these changes correlate with the severity of poisoning. Thirty patients with anticholinesterase pesticide poisoning were included, besides 10 healthy volunteers as a control group. Serum activities of butyrylcholinesterase (BuChE) and BG were measured for each subject on admission, then after 12 and 24 h. Serum levels of BuChE and BG in poisoned patients were significantly different from the control subjects; these differences persisted in repeated measurements. Moreover, the serum levels showed significant differences within each group of the three time points. A significant negative correlation was found between the serum activities of BuChE and BG in all groups at the three time points. In conclusion, serum BG activity seems a reliable marker for OP poisoning even when measured at 24 h after poisoning.

Computation-Aided Rational Deletion of C-Terminal Region Improved the Stability, Activity, and Expression Level of GH2 β-Glucuronidase.

In this study, computation-aided design on the basis of structural analysis was employed to rationally identify a highly dynamic C-terminal region that regulates the stability, expression level, and activity of a GH2 fungal glucuronidase from Aspergillus oryzae Li-3 (PGUS). Then, four mutants with a precisely truncated C-terminal region in different lengths were constructed; among them, mutant D591-604 with a 3.8-fold increase in half-life at 65 °C and a 6.8 kJ/mol increase in Gibbs free energy showed obviously improved kinetic and thermodynamic stability in comparison to PGUS. Mutants D590-604 and D591-604 both showed approximately 2.4-fold increases in the catalytic efficiency kcat/ Km and 1.8-fold increases in the expression level. Additionally, the expression level of PGUS was doubled through a C-terminal region swap with bacterial GUS from E. coli (EGUS). Finally, the robust PGUS mutants D590-604 and D591-604 were applied in the preparation of glycyrrhetinic acid with 4.0- and 4.4-fold increases in concentration through glycyrrhizin hydrolysis by a fed-batch process.

INCREASING NODULE SIZE1 Expression Is Required for Normal Rhizobial Symbiosis and Nodule Development.

Nodulation is crucial for biological nitrogen fixation (BNF) in legumes, but the molecular mechanisms underlying BNF have remained elusive. Here, we cloned a candidate gene underlying a major nodulation quantitative trait locus in soybean (Glycine max), INCREASING NODULE SIZE1 (GmINS1). GmINS1 encodes a cell wall β-expansin and is expressed primarily in vascular bundles, along with cortical and parenchyma cells of nodules. Four single-nucleotide polymorphisms distinguishing the two parents were found in the GmINS1 promoter region. Among them, single-nucleotide polymorphism A/C has a significant effect on GmINS1 expression in the parental genotype P2, based on β-glucuronidase activity and promoter deletion analysis. The expression of GmINS1 and the P2 genotype promoter was strongly associated with nodule development, not only in the parents but also in 40 progeny lines and 40 genotypes selected from a soybean core collection. Overexpression of GmINS1 resulted in increases in the number, biomass, infection cell abundance, and nitrogenase activity of large nodules and subsequently changed the nitrogen content and biomass of soybean plants. GmINS1 suppression via RNA interference had the opposite effect. Double suppression of GmEXPB2 and GmINS1 dramatically inhibited soybean nodulation. Our results reveal that GmINS1 is a critical gene in nodule development and that GmEXPB2 and GmINS1 synergistically control nodulation in soybean. Our findings shed light on the genetic basis of soybean nodulation and provide a candidate gene for optimizing BNF capacity through molecular breeding in soybean.

Comparison of Root Surface Enzyme Activity of Ericaceous Plants and Picea abies Growing at the Tree Line in the Austrian Alps

Plants with ericoid mycorrhizal and ectomycorrhizal associations coexist at the tree line and in many boreal forests. Both ericoid mycorrhizal and ectomycorrhizal roots are known to produce extracellular enzymes, but ericoid mycorrhizal fungi have been demonstrated in vitro to have higher enzyme activities. On hair roots of four ericoid mycorrhizal species (Rhododendron ferrugineum, Vaccinium vitis-idaea, Vaccinium myrtillus, Calluna vulgaris) and on ectomycorrhizal and non-mycorrhizal root tips of Picea abies growing at the tree line (1700 m) in the Austrian Alps, potential activities of eight extracellular root enzymes were estimated. Our results show that the activities of all the different extracellular root enzymes were generally similar among the ericaceous plant species. The mean laccase enzyme activity of ectomycorrhizal root tips of Picea abies was significantly higher than that of both the hair roots and fine roots of the ericaceous species. Leucine-amino-peptidase activity on hair roots was significantly higher than on non-mycorrhizal fine roots for the ericaceous vegetation. However, the mean activity of β-glucuronidase of the ericaceous species was significantly higher in fine roots compared to the hair roots. Generally extracellular root enzyme activity is not higher on ericaceous roots compared to ectomycorrhizas of Picea abies.

Stable transformation of Spirulina (Arthrospira) platensis: a promising microalga for production of edible vaccines.

The planktonic blue-green microalga Spirulina (Arthrospira) platensis possesses important features (e.g., high protein and vital lipids contents as well as essential vitamins) and can be consumed by humans and animals. Accordingly, this microalga gained growing attention as a new platform for producing edible-based pharmaceutical proteins. However, there are limited successful strategies for the transformation of S. platensis, in part because of an efficient expression of strong endonucleases in its cytoplasm. In the current work, as a pilot step for the expression of therapeutic proteins, an Agrobacterium-based system was established to transfer gfp:gus and hygromycin resistance (hygr) genes into the genome of S. platensis. The presence of acetosyringone in the transfection medium significantly reduced the transformation efficiency. The PCR and real-time RT-PCR data confirmed the successful integration and transcription of the genes. Flow cytometry and β-glucuronidase (GUS) activity experiments confirmed the successful production of GFP and the enzyme. Moreover, the western blot analysis showed a ~ 90kDa band in the transformed cells, indicating the successful production of the GFP:GUS protein. Three months after the transformation, the gene expression stability was validated by histochemical, flow cytometry, and hygromycin B resistance analyses.

Effective Treatment of Chronic Proliferative Cholangitis by Local Gentamicin Infusion in Rabbits.

Background Hepatolithiasis is highly prevalent in East Asia characterized by the presence of gallstones in the biliary ducts of the liver. Surgical resection is the potentially curative treatment but bears a high risk of stone recurrence and biliary restenosis. This is closely related to the universal presence of chronic proliferative cholangitis (CPC) in the majority of patients. Recent evidence has indicated the association of bacterial infection with the development of CPC in hepatolithiasis. Thus, this study aims to investigate the feasibility and efficacy of local infusion of gentamicin (an antibiotic) for the treatment of CPC in a rabbit model. Methods The rabbit CPC model was established based on previously published protocols. Bile duct samples were collected from gentamicin-treated or control animals for pathological and molecular characterization. Results Histologically, the hyperplasia of biliary epithelium and submucosal glands were inhibited and the thickness of the bile duct wall was significantly decreased after gentamicin therapy. Consistently, the percentage of proliferating cells marked by ki67 was significantly reduced by the treatment. More importantly, this treatment inhibited interleukin 2 production, an essential inflammatory cytokine, and the enzyme activity of endogenous β-Glucuronidase, a key factor in the formation of bile pigment. Conclusions Local gentamicin infusion effectively inhibits the inflammation, cell proliferation, and lithogenesis in a rabbit model of CPC. This approach represents a potential treatment for CPC and thus prevents recurrent hepatolithiasis.

Darunavir alleviates irinotecan-induced intestinal toxicity in Vivo

Irinotecan (CPT-11) is used to treat various cancers but side effects such as delayed diarrhea restrict its use. Darunavir (DRV) is an antiretroviral drug used to treat and prevent human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS), but whether DRV is protective against CPT-11-induced intestinal toxicity is unclear. An CPT-11-induced intestinal toxicity model was produced using uninterrupted CPT-11 (ip) for 4 d in mice. Enzyme-linked immuno sorbent assay (ELISA), fecal occult blood test (FOBT), Western blot, histopathological evaluation, and immunohistochemistry staining assays were used to document toxicity. DRV treatment attenuated CPT-11-induced intestinal toxicity via decreasing fecal occult blood and mitigating delayed-onset diarrhea, as well as reducing weight loss, reduced food intake, and pathomorphologic changes without inhibiting β-glucuronidase (β-GLU) activity. The high mobility group box-1 protein (HMGB1)-toll-like receptor 4 (TLR4) pathway induced inflammation and tight junction protein (occludin and zonular occluden-1) reduction in the colon was inhibited by DRV. Hepatotoxicity induced by CPT-11 was diminished after treatment with DRV, and activation of the NOD-like receptor 3 inflammasome (NLRP3) was prevented in colon tissue. In addition, DRV didn’t reduce the concentration of CPT-11 and 7-ethyl-10-hydroxycamptothecin (SN-38) in plasma at the same dose of irinotecan with DRV. DRV has anti-inflammatory and intestinal-protective properties and may be used to manage CPT-11-induced intestinal toxicity.

Serum Exoglycosidases in Children and Adolescents With Harmful Alcohol Use.

There is a lack of accurate alcohol-use biomarkers in children/adolescents due to a short drinking duration/rapid normalization of elevated markers. We checked if lysosomal exoglycosidases, elevated earlier in binge-drinking young adults, can be applicable in children/adolescents as markers of harmful alcohol use. The serum activities (pKat/mL) of α-fucosidase (FUC), β-galactosidase (GAL), β-glucuronidase (GLU), β-hexosaminidase (HEX; its HEX A and HEX B isoenzymes), and α-mannosidase (MAN) were determined in 20 healthy controls (C) and 25 children/adolescents with harmful alcohol use (intoxicated by alcohol at hospital admission -AI1 and on the next day -AI2). The serum HEX A and alanine aminotransferase (ALT) activity was significantly higher in the AI1 group than in the control. The activities of FUC, GAL, GLU, HEX B, and MAN were lower in the AI group. We found fair and poor accuracy, respectively, for increased enzymes HEX A and ALT. We found fair accuracy for decreased HEX B (AI1) and MAN (AI1), good accuracy for GLU (AI2), FUC (AI2), GAL (AI1, AI2), MAN (AI2), and excellent for FUC (AI1). Correlations were found: ALT with C-reactive protein (CRP), HEX A with white blood cell (WBC) count, blood alcohol concentration with FUC, MAN and HEX B, and WBC with FUC. Decreased FUC, GLU, GAL, MAN values, and especially FUC (AI1) have the potential to be markers of harmful alcohol use in children/adolescents. The raised activity of HEX A and ALT points to the need for further research to check another inflammatory agent as potential alcohol marker in children and adolescents. Samples need to be collected before intravenous fluid therapy.

Furan carbonylhydrazones-derived elicitors that induce the resistance of rice to the brown planthopper Nilaparvata lugens

To obtain new elicitors that could induce plant defense against pests, a series of furan carbonylhydrazones were synthesized. Phenotypic high-throughput screening of β-glucuronidase (GUS) activity was used to identify elicitors with strong activity. Integrated bioassays were conducted to acquire chemical elicitors that induce resistance in rice to the brown planthopper Nilaparvata lugens and they are not direct toxic to the pest. Our results demonstrate that these compounds have the potential to serve as new kinds of environment-friendly plant protection agents in rice.

Changes in hepatic metabolic enzyme activities and biliary excretion of 4-nitrophenol in streptozotocin induced diabetic rats

Activity of hepatic metabolic enzymes of glucuronidation and sulfation of 4-nitrophenol (PNP) and biliary excretion of its glucuronide (PNP-G) and sulfate (PNP-S) conjugates have been investigated in control and streptozotocin (STZ)-induced diabetic rats. 500 μM PNP solution was luminally perfused in a cannulated jejunal loop for 90 minutes. It was found that biliary excretion of PNP-G was significantly decreased in the diabetic rats. This effect of STZ could be completely reversed by administration of rapid-acting insulin. Activity of hepatic UDP-glucuronyltransferase and β-glucuronidase was also depressed by the STZ pretreatment. Administration of insulin antagonized the inhibitory action of STZ on UDP-glucuronyltransferase, but the reduced activity of β-glucuronidase was not reversed. Biliary excretion of PNP-S was also depressed in the diabetic rats. Whereas, different effects of insulin administration were observed. Namely, the lower biliary excretion rate of PNP-S was not changed after administration of insulin. Activity of the sulfotransferase and the arylsulfatase enzymes was not altered either by STZ pretreatment or by insulin administration. Biliary excretion of PNP was also significantly depressed by STZ and this depression was not changed after insulin administration. The results call attention to hepatobiliary circulation of low molecular weight xenobiotics and their glucuronide and sulfate conjugates.

Central role of the proximal tubular \u03b1Klotho/FGF receptor complex in FGF23-regulated phosphate and vitamin D metabolism

Fibroblast growth factor 23 (FGF23) plays critical roles in phosphate handling and vitamin D metabolism in the kidney. However, the effector cells of FGF23 in the kidney remain unclear. αKlotho, a putative enzyme possessing β-glucuronidase activity and also a permissive co-receptor for FGF23 to bind to FGF receptors (FGFRs), is expressed most abundantly in distal convoluted tubules, whereas it is expressed modestly in proximal tubules. Key molecular players of phosphate homeostasis and vitamin D-metabolizing enzymes are known to localize in proximal tubules. To clarify the direct function of FGF23 on proximal tubules, we ablated αKlotho or Fgfr1–4 genes specifically from these tubules using the Cre-loxP-mediated genetic recombination. Both conditional knockout mouse lines showed similar phenotypes that resembled those of systemic αKlotho or Fgf23 knockout mice. Compared with control mice, they showed significantly elevated levels of plasma phosphate, FGF23 and 1,25-dihydroxyvitamin D, ectopic calcification in the kidney and aging-related phenotypes like growth retardation, osteoporosis and shortened lifespan. These findings suggest that the primary function of FGF23 on mineral metabolism is mediated through αKlotho/FGFR co-receptors expressed in proximal tubular cells, and that the putative enzymatic function of αKlotho in the proximal tubule has a minor role in systemic mineral metabolism.

Effect of Saccharomyces Boulardii Cell Wall Extracts on Colon Cancer Prevention in Male F344 Rats Treated with 1,2-Dimethylhydrazine

ABSTRACTThe effect of Saccharomyces boulardii cell wall extracts on colon cancer prevention in rats treated with 1,2-dimethylhydrazine was investigated. A crude insoluble glucan (0.5 and 1.0 mg/kg/day) and a crude mannoprotein extract (0.3 and 3.0 mg/kg/day) were administered in rats by gavage for 12 weeks along with a high fat low fiber diet whereupon rats were sacrificed and aberrant crypt foci (ACF) were counted in the colon. Moreover, NAD(P)H: quinone reductase (QR) and harmful fecal enzymes (β-glucosidase and β-glucuronidase) were quantified in the liver and in the caecum, respectively. Results showed a reduction in ACF counts, a decreased β-glucuronidase activity and an increased QR activity when rats were treated only with insoluble glucan. While these enzymatic modulations may be constituted one of the mechanisms that is responsible for the reduction of ACF counts observed, the reduction of ACF counts caused by insoluble glucan should be addressed, at least, as a biomarker of their cancer-prevention properties. To our knowledge, this is the first study demonstrated that crude cell wall extract obtained from S. boulardii could have a potential role in colon cancer prevention in vivo by revealing the potential implication of QR and β-glucuronidase modulation.

Vascular preferential activity of the Pennisetum purpureum cinnamyl alcohol dehydrogenase promoter in transgenic tobacco plants

Little is known about the cross talk between the lignin biosynthesis gene promoters and the regulatory proteins that modulate molecular signaling and respond to various stresses. In this study, we characterized the promoter region of the lignin biosynthesis pathway cinnamyl alcohol dehydrogenase (CAD) gene in elephant grass, Pennisetum purpureum. Quantification of the transcript levels of the PpCAD promoter revealed it is preferentially expressed in vascular tissue, especially xylem. Histochemical and fluorometric assays confirmed the vascular-preferential expression of the PpCAD promoter, as the highest β-glucuronidase (GUS) activity was found in the basal stem in transgenic tobacco plants expressing a 1154-bp PpCAD promoter-GUS fusion construct. Moreover, 5′-deleted PpCAD promoter analyses showed that the 1154-bp PpCAD promoter fragment had the highest transcriptional activity, whereas the 2054-bp fragment had multifarious inducible activity responding to gibberellin (GA), methyl jasmonate (MeJA), abscisic acid (ABA), and wounding. The regions from −248 to −243 bp and −1416 to −1411 bp contained W-box cis-elements, which were detected by electrophoretic mobility shift assay (EMSA). The binding effects of the GA-responsive elements (from −561 to −555 bp and −1077 to −1071 bp), MeJA-responsive element (from −1146 to −1142 bp), and the ABA-responsive cis-element (from −1879 to −1874 bp) were also validated by EMSA. Based on our results, we suggest that lignin deposition associated with PpCAD promoter activity adapts to the environment through molecular signaling involving GA, MeJA, and ABA.

The Effect of Gut Bacterial β‐Glucuronidase on Serum Bilirubin Levels

The results of several large population studies have consistently demonstrated an inverse correlation between serum bilirubin levels and the development of cardiovascular and metabolic diseases. The goal of this study was to test the hypothesis that increased bacterial β‐Glucuronidase (β‐glu) activity can increase the levels of serum bilirubin by decreasing conjugation of bilirubin in the gut. Two strains of Lactobacillus acidophilus with different levels of β‐glu activity, NCK 56 (control) and NCK 2136 (constitutive overexpression), were grown and administered to mice once per day for five days. Each mouse was then administered 2 mL of fluid containing 500 million cells/ml via gastric gavage. Plasma samples were obtained prior to and after treatment. Serum bilirubin levels were measured using a specific assay kit via absorption at 700 nm. Serum bilirubin levels were similar in both groups prior to treatment averaging 2.3 +0.1 and 2.1 + 0.2 mg/dL in 56 and 2136 treated mice, respectively (n=3/group, p=0.8). Five day treatment of mice with control 56 L. acidophilus resulted in a significant decrease in serum bilirubin levels to 1.7 + 0.5 mg/dL (p&lt;0.05), while treatment with β‐glu overexpressing L. acidophilus did not change serum bilirubin levels which averaged 2.3 + 0.3 mg/dL. Our results demonstrate that β‐Glu expression by modified L. acidophilus prevented the decrease in bilirubin that is seen upon administration of the control L. acidophilus bacterial strain. Overexpression β‐glu in a different host bacteria species with less reducing activity may provide a way to elevate serum bilirubin levels.Support or Funding InformationHearin Support Foundation, Medical Student Research Program, University of Mississippi Medical Center School of Medicine. National Heart, Lung, and Blood Institute, IP01HL‐051971. National Institute of General Medical Sciences, P20GM‐104357.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Design, synthesis and in vitro evaluation of β-glucuronidase-sensitive prodrug of 5-aminolevulinic acid for photodiagnosis of breast cancer cells

Treatment of cancer cells by clinically approved hexyl ester of 5-aminolevulinic acid (ALA-Hex) induces accumulation of fluorescent porphyrins in tumors. This allows fluorescence photodiagnosis (PD) of bladder cancer by blue light illumination. However, PD of other cancers is hampered by acute toxicity of the compound limiting its use to local applications. We have designed and synthesized a new prodrug of ALA-Hex that tackles the stability-activity paradox of amino-modified 5-ALA prodrugs. The glucuronide prodrug Glu-ALA-Hex demonstrates excellent stability under physiological conditions and activation in the presence of the target enzyme. β-glucuronidase-triggered release of 5-ALA is programmed to yield fluorescence in tumor environment with elevated β-glucuronidase activity, a characteristic of many solid tumors. Glu-ALA-Hex produces similar levels of fluorescence as ALA-Hex in breast cancer MCF7 cells in vitro but with much lower non-specific cell toxicity.