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1.
Glycoconj J ; 2024 Jul 02.
Artículo en Inglés | MEDLINE | ID: mdl-38954268

RESUMEN

A glucosyl-rich pectin, JMMP-3 (Mw, 2.572 × 104 g/mol, O-methyl % = 3.62%), was isolated and purified from the pericarp of the immature fruit of Juglans mandshurica Maxim. (QingLongYi). The structure of JMMP-3 was studied systematically by infrared spectroscopy, monosaccharide compositions, methylation analysis, partial acid hydrolysis, and 1/2D-NMR. The backbone of JMMP-3 possessed a smooth region (→ 4GalA1 →) and a hairy region (→ 4GalA1 → 2Rha1 →) with a molar ratio of 2: 5. The substitution of four characteristic side chains (R1-R4) occurs at C-4 of → 2,4)-α-Rhap-(1→, where R1 is composed of → 5)-α-Araf-(1→, R2 is composed of → 4)-ß-Galp-(1 → and ß-Galp-(1→, R3 is composed of α-Glcp-(1→, →4)-α-Glcp-(1 → and → 4,6)-α-Glcp-(1→, and R4 is composed of → 5)-α-Araf-(1→, ß-Galp-(1→, → 4)-ß-Galp-(1→, → 3,4)-ß-Galp-(1→, → 4,6)-ß-Galp-(1 → and → 2,4)-ß-Galp-(1 → . In addition, the antitumor activity of JMMP-3 on HepG2 cells was preliminarily investigated.

2.
Drug Metab Dispos ; 52(7): 681-689, 2024 Jun 17.
Artículo en Inglés | MEDLINE | ID: mdl-38719743

RESUMEN

This study aimed to characterize the effects of arsenic exposure on the expression of microsomal epoxide hydrolase (mEH or EPHX1) and soluble epoxide hydrolase (sEH or EPHX2) in the liver and small intestine. C57BL/6 mice were exposed to sodium arsenite in drinking water at various doses for up to 28 days. Intestinal, but not hepatic, mEH mRNA and protein expression was induced by arsenic at 25 ppm, in both males and females, whereas hepatic mEH expression was induced by arsenic at 50 or 100 ppm. The induction of mEH was gene specific, as the arsenic exposure did not induce sEH expression in either tissue. Within the small intestine, mEH expression was induced only in the proximal, but not the distal segments. The induction of intestinal mEH was accompanied by increases in microsomal enzymatic activities toward a model mEH substrate, cis-stilbene oxide, and an epoxide-containing drug, oprozomib, in vitro, and by increases in the levels of PR-176, the main hydrolysis metabolite of oprozomib, in the proximal small intestine of oprozomib-treated mice. These findings suggest that intestinal mEH, playing a major role in converting xenobiotic epoxides to less reactive diols, but not sEH, preferring endogenous epoxides as substrates, is relevant to the adverse effects of arsenic exposure, and that further studies of the interactions between drinking water arsenic exposure and the disposition or possible adverse effects of epoxide-containing drugs and other xenobiotic compounds in the intestine are warranted. SIGNIFICANCE STATEMENT: Consumption of arsenic-contaminated water has been associated with increased risks of various adverse health effects, such as diabetes, in humans. The small intestinal epithelial cells are the main site of absorption of ingested arsenic, but they are not well characterized for arsenic exposure-related changes. This study identified gene expression changes in the small intestine that may be mechanistically linked to the adverse effects of arsenic exposure and possible interactions between arsenic ingestion and the pharmacokinetics of epoxide-containing drugs in vivo.


Asunto(s)
Agua Potable , Epóxido Hidrolasas , Intestino Delgado , Ratones Endogámicos C57BL , Animales , Epóxido Hidrolasas/metabolismo , Epóxido Hidrolasas/genética , Ratones , Masculino , Femenino , Intestino Delgado/efectos de los fármacos , Intestino Delgado/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/enzimología , Arsénico/toxicidad , Arsénico/metabolismo , Arsenitos/toxicidad , Arsenitos/metabolismo , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efectos de los fármacos , Microsomas/efectos de los fármacos , Microsomas/metabolismo , Microsomas/enzimología , Compuestos de Sodio/toxicidad
3.
Toxicol Lett ; 397: 89-102, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38768835

RESUMEN

Aging increases susceptibility to lung disease, but the topic is understudied, especially in relation to environmental exposures with the bulk of rodent studies using young adults. This study aims to define the pulmonary toxicity of naphthalene (NA) and the impacts of a dietary antioxidant, ergothioneine (ET), in the liver and lungs of middle-aged mice. NA causes a well-characterized pattern of conducting airway epithelial injury in the lung in young adult mice, but NA's toxicity has not been characterized in middle-aged mice, aged 1-1.5 years. ET is a dietary antioxidant that is synthesized by bacteria and fungi. The ET transporter (ETT), SLC22A4, is upregulated in tissues that experience high levels of oxidative stress. In this study, middle-aged male and female C57BL/6 J mice, maintained on an ET-free synthetic diet from conception, were gavaged with 70 mg/kg of ET for five consecutive days. On day 8, the mice were exposed to a single intraperitoneal NA dose of 50, 100, 150, or 200 mg/kg. At 24 hours post NA injection samples were collected and analyzed for ET concentration and reduced (GSH) and oxidized glutathione (GSSG) concentrations. Histopathology, morphometry, and gene expression were examined. Histopathology of mice exposed to 100 mg/kg of NA suggests reduction in toxicity in the terminal airways of both male (p ≤ 0.001) and female (p ≤ 0.05) middle-aged mice by the ET pretreatment. Our findings in this study are the first to document the toxicity of NA in middle-aged mice and show some efficacy of ET in reducing NA toxicity.


Asunto(s)
Envejecimiento , Antioxidantes , Ergotioneína , Pulmón , Naftalenos , Ergotioneína/uso terapéutico , Naftalenos/toxicidad , Pulmón/patología , Pulmón/fisiología , Humanos , Suplementos Dietéticos , Masculino , Femenino , Animales , Ratones , Antioxidantes/uso terapéutico , Reacción en Cadena de la Polimerasa , Expresión Génica , Glutatión/genética , Glutatión/metabolismo
4.
J Med Chem ; 67(9): 7301-7311, 2024 May 09.
Artículo en Inglés | MEDLINE | ID: mdl-38635879

RESUMEN

Although the selective and effective clearance of senescent cancer cells can improve cancer treatment, their development is confronted by many challenges. As part of efforts designed to overcome these problems, prodrugs, whose design is based on senescence-associated ß-galactosidase (SA-ß-gal), have been developed to selectively eliminate senescent cells. However, chemotherapies relying on targeted molecular inhibitors as senolytic drugs can induce drug resistance. In the current investigation, we devised a new strategy for selective degradation of target proteins in senescent cancer cells that utilizes a prodrug composed of the SA-ß-gal substrate galactose (galacto) and the proteolysis-targeting chimeras (PROTACs) as senolytic agents. Prodrugs Gal-ARV-771 and Gal-MS99 were found to display senolytic indexes higher than those of ARV-771 and MS99. Significantly, results of in vivo studies utilizing a human lung A549 xenograft mouse model demonstrated that concomitant treatment with etoposide and Gal-ARV-771 leads to a significant inhibition of tumor growth without eliciting significant toxicity.


Asunto(s)
Senescencia Celular , Galactosa , Profármacos , Proteolisis , Humanos , Animales , Senescencia Celular/efectos de los fármacos , Galactosa/química , Galactosa/farmacología , Profármacos/farmacología , Profármacos/química , Profármacos/uso terapéutico , Ratones , Proteolisis/efectos de los fármacos , Antineoplásicos/farmacología , Antineoplásicos/química , Antineoplásicos/síntesis química , Antineoplásicos/uso terapéutico , Ensayos Antitumor por Modelo de Xenoinjerto , beta-Galactosidasa/metabolismo , Ratones Desnudos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Células A549 , Etopósido/farmacología , Senoterapéuticos/farmacología , Senoterapéuticos/química , Quimera Dirigida a la Proteólisis
5.
Nat Commun ; 15(1): 2073, 2024 Mar 07.
Artículo en Inglés | MEDLINE | ID: mdl-38453918

RESUMEN

Cholesterol (Chol) fortifies packing and reduces fluidity and permeability of the lipid bilayer in vesicles (liposomes)-mediated drug delivery. However, under the physiological environment, Chol is rapidly extracted from the lipid bilayer by biomembranes, which jeopardizes membrane stability and results in premature leakage for delivered payloads, yielding suboptimal clinic efficacy. Herein, we report a Chol-modified sphingomyelin (SM) lipid bilayer via covalently conjugating Chol to SM (SM-Chol), which retains membrane condensing ability of Chol. Systemic structure activity relationship screening demonstrates that SM-Chol with a disulfide bond and longer linker outperforms other counterparts and conventional phospholipids/Chol mixture systems on blocking Chol transfer and payload leakage, increases maximum tolerated dose of vincristine while reducing systemic toxicities, improves pharmacokinetics and tumor delivery efficiency, and enhances antitumor efficacy in SU-DHL-4 diffuse large B-cell lymphoma xenograft model in female mice. Furthermore, SM-Chol improves therapeutic delivery of structurally diversified therapeutic agents (irinotecan, doxorubicin, dexamethasone) or siRNA targeting multi-drug resistant gene (p-glycoprotein) in late-stage metastatic orthotopic KPC-Luc pancreas cancer, 4T1-Luc2 triple negative breast cancer, lung inflammation, and CT26 colorectal cancer animal models in female mice compared to respective FDA-approved nanotherapeutics or lipid compositions. Thus, SM-Chol represents a promising platform for universal and improved drug delivery.


Asunto(s)
Membrana Dobles de Lípidos , Esfingomielinas , Humanos , Femenino , Ratones , Animales , Membrana Dobles de Lípidos/química , Esfingomielinas/química , Liposomas/química , Fosfolípidos/química , Colesterol/química
6.
J Pharm Pharmacol ; 76(4): 307-326, 2024 Apr 03.
Artículo en Inglés | MEDLINE | ID: mdl-38243389

RESUMEN

OBJECTIVES: Oligo-/polysaccharides from Cyathula officinalis Kuan (COPs) and Achyranthes bidentata Blume (ABPs) have attracted researchers' attention in the fields of healthy food supplements and traditional Chinese medicine (Niúxi) due to their multiple bioactivities combined with their nontoxic and highly biocompatible nature. The purpose of this paper was to provide a systematic and comprehensive overview of the extraction, purification, and structural analysis methods, chemical characteristics, biological activities, and structure bioactivity relationship. Furthermore, the possible development trends and perspectives for future research, and traditional uses of Niúxi are also summarized. METHODS: All the information was gathered from a library search and scientific databases. KEY FINDINGS: Although COPs and ABPs are derived from different plants, they have similar structural features in type, structure, and glycosidic linkage patterns and biological activities in vivo and in vitro. However, there are differences in monosaccharide compositions, which can be used as an identification mark. CONCLUSIONS: As traditional Chinese herbal medicine, C. officinalis and A. bidentata have similar pharmacological activities. The COPs and ABP possess wide pharmacological effects such as antitumor, antioxidant, anti-osteoporosis, and anti-inflammatory. Meanwhile, the biological activity and structure-activity relationship of purified COPs and ABPs are less studied, future research should focus on them.


Asunto(s)
Achyranthes , Amaranthaceae , Osteoporosis , Achyranthes/química , Polisacáridos/farmacología , Polisacáridos/química
7.
Mol Neurobiol ; 61(4): 1892-1906, 2024 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-37814108

RESUMEN

Alzheimer's disease (AD) is the most common chronic progressive neurodegenerative disease in the elderly. It has an increasing prevalence and a growing health burden. One of the limitations in studying AD is the lack of animal models that show features of Alzheimer's pathogenesis. The tree shrew has a much closer genetic affinity to primates than to rodents and has great potential to be used for research into aging and AD. In this study, we aimed to investigate whether tree shrews naturally develop cognitive impairment and major AD-like pathologies with increasing age. Pole-board and novel object recognition tests were used to assess the cognitive performance of adult (about 1 year old) and aged (6 years old or older) tree shrews. The main AD-like pathologies were assessed by Western blotting, immunohistochemical staining, immunofluorescence staining, and Nissl staining. Our results showed that the aged tree shrews developed an impaired cognitive performance compared to the adult tree shrews. Moreover, the aged tree shrews exhibited several age-related phenotypes that are associated with AD, including increased levels of amyloid-ß (Aß) accumulation and phosphorylated tau protein, synaptic and neuronal loss, and reactive gliosis in the cortex and the hippocampal tissues. Our study provides further evidence that the tree shrew is a promising model for the study of aging and AD.


Asunto(s)
Enfermedad de Alzheimer , Disfunción Cognitiva , Enfermedades Neurodegenerativas , Anciano , Animales , Humanos , Niño , Lactante , Enfermedad de Alzheimer/patología , Tupaia/metabolismo , Tupaiidae/metabolismo , Musarañas/metabolismo , Disfunción Cognitiva/metabolismo , Proteínas tau/genética , Péptidos beta-Amiloides/metabolismo , Modelos Animales de Enfermedad , Cognición
8.
Eur J Med Chem ; 264: 116039, 2024 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-38103540

RESUMEN

P-glycoprotein (P-gp) is an important factor leading to multidrug resistance (MDR) in cancer treatment. The co-administration of anticancer drugs and P-gp inhibitors has been a treatment strategy to overcome MDR. In recent years, tyrosine kinase inhibitor Lapatinib has been reported to reverse MDR through directly interacting with ABC transporters. In this work, a series of P-gp inhibitors (1-26) was designed and synthesized by integrating the quinazoline core of Lapatinib into the molecule framework of the third-generation P-gp inhibitor Tariquidar. Among them, compound 14 exhibited better MDR reversal activity than Tariquidar. The docking results showed compound 14 displayed the L-shaped molecular conformation. Importantly, compound 14 increased the accumulation of Adriamycin (ADM) and rhodamine 123 (Rh123) in MCF7/ADM cells. Besides, compound 14 significantly increased ADM-induced apoptosis and inhibited the proliferation, migration and invasion of MCF7/ADM cells. It was also demonstrated that compound 14 significantly inhibited the growth of MCF7/ADM xenograft tumors by increasing the sensitivity of ADM. In summary, compound 14 has the potential to overcome MDR caused by P-gp.


Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP , Antineoplásicos , Humanos , Lapatinib , Resistencia a Antineoplásicos , Resistencia a Múltiples Medicamentos , Antineoplásicos/farmacología , Subfamilia B de Transportador de Casetes de Unión a ATP , Doxorrubicina/farmacología , Benzamidas/farmacología
9.
Zhongguo Zhong Yao Za Zhi ; 48(18): 4874-4883, 2023 Sep.
Artículo en Chino | MEDLINE | ID: mdl-37802829

RESUMEN

Rheumatoid arthritis(RA) is a widely prevalent autoimmune inflammatory disease that severely affects patients' quality of life. Currently, conventional formulations against RA have several limitations, such as nonspecificity, poor efficacy, large drug dosages, frequent administration, and systemic side effects. Nanotechnology-based drug delivery systems have emerged as a promising stra-tegy for the diagnosis and treatment of RA since nanotechnology can overcome the limitations of traditional treatments and simplify the complexity of the disease. These systems enable targeted delivery of anti-inflammatory drugs to the inflamed areas through active and passive targeting, achieving specificity to the joints, overcoming the need for increased dosage and administration frequency, and reducing associated adverse reactions. This article aimed to review nanocarrier-based drug delivery systems in the field of RA and elucidate how nanosystems can be utilized to deliver therapeutic drugs to inflamed joints for controlling RA progression. By discussing the current issues and challenges faced by nanodrug delivery systems and highlighting the urgent need for solutions, this article offers theoretical support for further research on nanotechnology-based co-delivery systems in the future.


Asunto(s)
Artritis Reumatoide , Enfermedades Autoinmunes , Humanos , Calidad de Vida , Sistemas de Liberación de Medicamentos , Artritis Reumatoide/tratamiento farmacológico , Enfermedades Autoinmunes/tratamiento farmacológico , Nanotecnología
10.
Chem Res Toxicol ; 36(8): 1386-1397, 2023 08 21.
Artículo en Inglés | MEDLINE | ID: mdl-37467352

RESUMEN

Polychlorinated biphenyls (PCBs) are environmental contaminants that can cause neurotoxicity. PCBs, such as PCB 95 (2,2',3,5',6-pentachlorobiphenyl), can be metabolized by cytochrome P450 enzymes into neurotoxic metabolites. To better understand how the metabolism of PCB 95 affects neurotoxic outcomes, we conducted a study on the disposition of PCB 95 in transgenic mouse models. The mice were given a single oral dose of PCB 95 (1.0 mg/kg) and were euthanized 24 h later for analysis. PCB 95 levels were highest in adipose tissue, followed by the liver, brain, and blood. Adipose tissue levels were significantly higher in wild-type (WT) mice than in Cyp2abfgs-null (KO) or CYP2A6-transgenic (KI) mice. We also observed genotype-dependent differences in the enrichment of aS-PCB 95 in female mice, with a less pronounced enrichment in KO than WT and KI mice. Ten hydroxylated PCB 95 metabolites were detected in blood and tissue across all exposure groups. The metabolite profiles differed across tissues, while sex and genotype-dependent differences were less pronounced. Total OH-PCB levels were highest in the blood, followed by the liver, adipose tissue, and brain. Total OH-PCB blood levels were lower in KO than in WT mice, while the opposite trend was observed in the liver. In male mice, total OH-PCB metabolite levels were significantly lower in KI than in WT mice in blood and the liver, while the opposite trend was observed in female mice. In conclusion, the study highlights the differences in the atropselective disposition of PCB 95 and its metabolites in different types of mice, demonstrating the usefulness of these transgenic mouse models for characterizing the role of PCB metabolism in PCB neurotoxicity.


Asunto(s)
Bifenilos Policlorados , Ratones , Masculino , Femenino , Animales , Bifenilos Policlorados/toxicidad , Sistema Enzimático del Citocromo P-450/genética , Sistema Enzimático del Citocromo P-450/metabolismo , Hígado/metabolismo , Hidroxilación , Ratones Transgénicos
11.
Mil Med Res ; 10(1): 31, 2023 Jul 14.
Artículo en Inglés | MEDLINE | ID: mdl-37443101

RESUMEN

Rheumatoid arthritis (RA) is a systemic autoimmune disease that is primarily manifested as synovitis and polyarticular opacity and typically leads to serious joint damage and irreversible disability, thus adversely affecting locomotion ability and life quality. Consequently, good prognosis heavily relies on the early diagnosis and effective therapeutic monitoring of RA. Activatable fluorescent probes play vital roles in the detection and imaging of biomarkers for disease diagnosis and in vivo imaging. Herein, we review the fluorescent probes developed for the detection and imaging of RA biomarkers, namely reactive oxygen/nitrogen species (hypochlorous acid, peroxynitrite, hydroxyl radical, nitroxyl), pH, and cysteine, and address the related challenges and prospects to inspire the design of novel fluorescent probes and the improvement of their performance in RA studies.


Asunto(s)
Artritis Reumatoide , Sinovitis , Humanos , Colorantes Fluorescentes/uso terapéutico , Artritis Reumatoide/diagnóstico por imagen , Artritis Reumatoide/tratamiento farmacológico , Sinovitis/diagnóstico , Biomarcadores , Especies de Nitrógeno Reactivo/uso terapéutico , Especies Reactivas de Oxígeno
13.
Emerg Microbes Infect ; 12(1): 2204164, 2023 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-37060263

RESUMEN

SARS-CoV-2 has caused a global pandemic with significant humanity and economic loss since 2020. Currently, only limited options are available to treat SARS-CoV-2 infections for vulnerable populations. In this study, we report a universal fluorescence polarization (FP)-based high throughput screening (HTS) assay for SAM-dependent viral methyltransferases (MTases), using a fluorescent SAM-analogue, FL-NAH. We performed the assay against a reference MTase, NSP14, an essential enzyme for SARS-CoV-2 to methylate the N7 position of viral 5'-RNA guanine cap. The assay is universal and suitable for any SAM-dependent viral MTases such as the SARS-CoV-2 NSP16/NSP10 MTase complex and the NS5 MTase of Zika virus (ZIKV). Pilot screening demonstrated that the HTS assay was very robust and identified two candidate inhibitors, NSC 111552 and 288387. The two compounds inhibited the FL-NAH binding to the NSP14 MTase with low micromolar IC50. We used three functional MTase assays to unambiguously verified the inhibitory potency of these molecules for the NSP14 N7-MTase function. Binding studies indicated that these molecules are bound directly to the NSP14 MTase with similar low micromolar affinity. Moreover, we further demonstrated that these molecules significantly inhibited the SARS-CoV-2 replication in cell-based assays at concentrations not causing cytotoxicity. Furthermore, NSC111552 significantly synergized with known SARS-CoV-2 drugs including nirmatrelvir and remdesivir. Finally, docking suggested that these molecules bind specifically to the SAM-binding site on the NSP14 MTase. Overall, these molecules represent novel and promising candidates to further develop broad-spectrum inhibitors for the management of viral infections.


Asunto(s)
COVID-19 , Infección por el Virus Zika , Virus Zika , Humanos , Metiltransferasas/genética , Metiltransferasas/metabolismo , SARS-CoV-2/genética , Ensayos Analíticos de Alto Rendimiento , Proteínas no Estructurales Virales/metabolismo , Virus Zika/genética , Virus Zika/metabolismo , Sitios de Unión , Caperuzas de ARN/química , Caperuzas de ARN/genética , Caperuzas de ARN/metabolismo , Polarización de Fluorescencia , ARN Viral/genética
14.
Bioorg Chem ; 133: 106382, 2023 04.
Artículo en Inglés | MEDLINE | ID: mdl-36716580

RESUMEN

Acute pancreatitis (AP) is a frequent abdominal inflammatory disease. Despite the high morbidity and mortality, the management of AP remains unsatisfactory. Disulfiram (DSF) is an FDA-proved drug with potential therapeutic effects on inflammatory diseases. In this study, we aim to investigate the effect of DSF on pancreatic acinar cell necrosis, and to explore the underlying mechanisms. Cell necrosis was induced by sodium taurocholate or caerulein, AP mice model was induced by nine hourly injections of caerulein. Network pharmacology, molecular docking, and molecular dynamics simulation were used to explore the potential targets of DSF in protecting against cell necrosis. The results indicated that DSF significantly inhibited acinar cell necrosis as evidenced by a decreased ratio of necrotic cells in the pancreas. Network pharmacology, molecular docking, and molecular dynamics simulation identified RIPK1 as a potent target of DSF in protecting against acinar cell necrosis. qRT-PCR analysis revealed that DSF decreased the mRNA levels of RIPK1 in freshly isolated pancreatic acinar cells and the pancreas of AP mice. Western blot showed that DSF treatment decreased the expressions of RIPK1 and MLKL proteins. Moreover, DSF inhibited NF-κB activation in acini. It also decreased the protein expression of TLR4 and the formation of neutrophils extracellular traps (NETs) induced by damage-associated molecular patterns released by necrotic acinar cells. Collectively, DSF could ameliorate the severity of mouse acute pancreatitis by inhibiting RIPK-dependent acinar cell necrosis and the following formation of NETs.


Asunto(s)
Pancreatitis , Ratones , Animales , Pancreatitis/tratamiento farmacológico , Pancreatitis/inducido químicamente , Células Acinares , Disulfiram/efectos adversos , Ceruletida/efectos adversos , Enfermedad Aguda , Simulación del Acoplamiento Molecular , Necrosis , Proteína Serina-Treonina Quinasas de Interacción con Receptores/farmacología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/uso terapéutico
15.
Drug Metab Dispos ; 51(1): 46-53, 2023 01.
Artículo en Inglés | MEDLINE | ID: mdl-36273825

RESUMEN

Most transgenic mouse models are generated through random integration of the transgene. The location of the transgene provides valuable information for assessing potential effects of the transgenesis on the host and for designing genotyping protocols that can amplify across the integration site, but it is challenging to identify. Here, we report the successful utility of optical genome mapping technology to identify the transgene insertion site in a CYP2A13/2B6/2F1-transgenic mouse model, which produces three human cytochrome P450 (P450) enzymes (CYP2A13, CYP2B6, and CYP2F1) that are encoded by neighboring genes on human chromosome 19. These enzymes metabolize many drugs, respiratory toxicants, and chemical carcinogens. Initial efforts to identify candidate insertion sites by whole genome sequencing was unsuccessful, apparently because the transgene is located in a region of the mouse genome that contains highly repetitive sequences. Subsequent utility of the optical genome mapping approach, which compares genome-wide marker distribution between the transgenic mouse genome and a reference mouse (GRCm38) or human (GRCh38) genome, localized the insertion site to mouse chromosome 14, between two marker positions at 4451324 base pair and 4485032 base pair. A transgene-mouse genome junction sequence was further identified through long-polymerase chain reaction amplification and DNA sequencing at GRCm38 Chr.14:4484726. The transgene insertion (∼2.4 megabase pair) contained 5-7 copies of the human transgenes, which replaced a 26.9-33.4 kilobase pair mouse genomic region, including exons 1-4 of Gm3182, a predicted and highly redundant gene. Finally, the sequencing results enabled the design of a new genotyping protocol that can distinguish between hemizygous and homozygous CYP2A13/2B6/2F1-transgenic mice. SIGNIFICANCE STATEMENT: This study characterizes the genomic structure of, and provides a new genotyping method for, a transgenic mouse model that expresses three human P450 enzymes, CYP2A13, CYP2B6, and CYP2F1, that are important in xenobiotic metabolism and toxicity. The demonstrated success in applying the optical genome mapping technology for identification of transgene insertion sites should encourage others to do the same for other transgenic models generated through random integration, including most of the currently available human P450 transgenic mouse models.


Asunto(s)
Hidrocarburo de Aril Hidroxilasas , Sistema Enzimático del Citocromo P-450 , Ratones , Animales , Humanos , Ratones Transgénicos , Citocromo P-450 CYP2B6/genética , Sistema Enzimático del Citocromo P-450/genética , Transgenes/genética , Modelos Animales de Enfermedad , Mapeo Cromosómico/métodos , Hidrocarburo de Aril Hidroxilasas/genética
16.
Chem Res Toxicol ; 35(12): 2310-2323, 2022 12 19.
Artículo en Inglés | MEDLINE | ID: mdl-36473170

RESUMEN

Chiral polychlorinated biphenyls (PCB) are environmentally relevant developmental neurotoxicants. Because their hydroxylated metabolites (OH-PCBs) are also neurotoxic, it is necessary to determine how PCB metabolism affects the developing brain, for example, in mouse models. Because the cytochrome P450 isoforms involved in the metabolism of chiral PCBs remain unexplored, we investigated the metabolism of PCB 91 (2,2',3,4',6-pentachlorobiphenyl), PCB 95 (2,2',3,5',6-pentachlorobiphenyl), PCB 132 (2,2',3,3',4,6'-hexachlorobiphenyl), and PCB 136 (2,2',3,3',6,6'-hexachlorobiphenyl) using liver microsomes from male and female Cyp2a(4/5)bgs-null, Cyp2f2-null, and wild-type mice. Microsomes, pooled by sex, were incubated with 50 µM PCB for 30 min, and the levels and enantiomeric fractions of the OH-PCBs were determined gas chromatographically. All four PCB congeners appear to be atropselectively metabolized by CYP2A(4/5)BGS and CYP2F2 enzymes in a congener- and sex-dependent manner. The OH-PCB metabolite profiles of PCB 91 and PCB 132, PCB congeners with one para-chlorine substituent, differed between null and wild-type mice. No differences in the metabolite profiles were observed for PCB 95 and PCB 136, PCB congeners without a para-chlorine group. These findings suggest that Cyp2a(4/5)bgs-null and Cyp2f2-null mice can be used to study how a loss of a specific metabolic function (e.g., deletion of Cyp2a(4/5)bgs or Cyp2f2) affects the toxicity of chiral PCB congeners.


Asunto(s)
Bifenilos Policlorados , Masculino , Femenino , Ratones , Animales , Bifenilos Policlorados/metabolismo , Microsomas Hepáticos/metabolismo , Familia 2 del Citocromo P450/metabolismo , Ratones Transgénicos , Cloro/metabolismo , Hidroxilación , Ratones Noqueados
17.
Antioxidants (Basel) ; 11(9)2022 Sep 18.
Artículo en Inglés | MEDLINE | ID: mdl-36139908

RESUMEN

Arsenic exposure has been associated with the risks of various diseases, including cancers and metabolic diseases. The aim of this study was to examine the effects of arsenic exposure via drinking water on the expression of heme oxygenase-1 (HO-1), a major responsive gene to arsenic-induced oxidative stress, in mouse intestinal epithelial cells which is the first site of exposure for ingested arsenic, and the liver, a known target of arsenic toxicity. The expression of HO-1 was determined at mRNA, protein, or enzymic activity levels in mice exposed to sodium arsenite through drinking water, at various doses (0, 2.5, 10, 25, 100 ppm), and for various time periods (1, 3, 7, or 28 days). HO-1 was significantly induced in the intestine, but not liver, at arsenic doses of 25 ppm or lower. The intestinal HO-1 induction was seen in both males and females, plateaued within 1-3 days of exposure, and was accompanied by increases in microsomal HO activity. In mice exposed to 25-ppm of arsenite for 7 days, total arsenic and As(III) levels in intestinal epithelial cells were significantly higher than in the liver. These findings identify intestinal epithelial cells as likely preferential targets for arsenic toxicity and support further studies on the functional consequences of intestinal HO-1 induction.

18.
ACS Infect Dis ; 8(9): 1851-1868, 2022 09 09.
Artículo en Inglés | MEDLINE | ID: mdl-35948057

RESUMEN

Drug resistance is a significant concern in the treatment of diseases, including cryptococcosis caused by Cryptococcus neoformans (Cne) and Cryptococcus gattii (Cga). Alternative drug targets are necessary to overcome drug resistance before it attains a critical stage. Splicing of inteins from pro-protein precursors is crucial for activities of essential proteins hosting intein elements in many organisms, including human pathogens such as Cne and Cga. Through a high-throughput screening, we identified calcimycin (CMN) as a potent Prp8 intein splicing inhibitor with a minimum inhibitory concentration (MIC) of 1.5 µg/mL against the wild-type Cne-H99 (Cne-WT or Cne). In contrast, CMN inhibited the intein-less mutant strain (Cne-Mut) with a 16-fold higher MIC. Interestingly, Aspergillus fumigatus and a few Candida species were resistant to CMN. Further studies indicated that CMN reduced virulence factors such as urease activity, melanin production, and biofilm formation in Cne. CMN also inhibited Cne intracellular infection in macrophages. In a target-specific split nanoluciferase assay, the IC50 of CMN was 4.6 µg/mL. Binding of CMN to recombinant Prp8 intein was demonstrated by thermal shift assay and microscale thermophoresis. Treating Cne cells with CMN reduced intein splicing. CMN was fungistatic and showed a synergistic effect with the known antifungal drug amphotericin B. Finally, CMN treatment at 20 mg/kg body weight led to 60% reduction in lung fungal load in a cryptococcal pulmonary infection mouse model. Overall, CMN represents a potent antifungal with a novel mechanism of action to treat Cne and possibly Cga infections.


Asunto(s)
Criptococosis , Cryptococcus neoformans , Animales , Antifúngicos/metabolismo , Antifúngicos/farmacología , Calcimicina/metabolismo , Calcimicina/farmacología , Criptococosis/tratamiento farmacológico , Proteínas Fúngicas/química , Humanos , Inteínas , Ratones , Alineación de Secuencia
19.
Acta Pharm Sin B ; 12(4): 1662-1670, 2022 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-35847519

RESUMEN

Zika virus (ZIKV) causes significant human diseases without specific therapy. Previously we found erythrosin B, an FDA-approved food additive, inhibited viral NS2B-NS3 interactions, leading to inhibition of ZIKV infection in cell culture. In this study, we performed pharmacokinetic and in vivo studies to demonstrate the efficacy of erythrosin B against ZIKV in 3D mini-brain organoid and mouse models. Our results showed that erythrosin B is very effective in abolishing ZIKV replication in the 3D organoid model. Although pharmacokinetics studies indicated that erythrosin B had a low absorption profile, mice challenged by a lethal dose of ZIKV showed a significantly improved survival rate upon oral administration of erythrosin B, compared to vehicle control. Limited structure-activity relationship studies indicated that most analogs of erythrosin B with modifications on the xanthene ring led to loss or reduction of inhibitory activities towards viral NS2B-NS3 interactions, protease activity and antiviral efficacy. In contrast, introducing chlorine substitutions on the isobenzofuran ring led to slightly increased activities, suggesting that the isobenzofuran ring is well tolerated for modifications. Cytotoxicity studies indicated that all derivatives are nontoxic to human cells. Overall, our studies demonstrated erythrosin B is an effective antiviral against ZIKV both in vitro and in vivo.

20.
Front Med (Lausanne) ; 9: 876207, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35573022

RESUMEN

Background: Bloodstream infections (BSI) are one of the common causes of morbidity and mortality in hospitals; however, the pathogenic spectrum and bacterial antibiotic resistance vary across the world. Therefore, identifying the pathogenic spectrum and changes in bacterial antibiotic resistance is critical in controlling BSI and preventing the irrational use of antibiotics. This study evaluated the microbiological and clinical data of BSI patients in the intensive care unit (ICU) of Tianjin Medical University General Hospital in Tianjin, China, to guide the selection of empirical antibiotic therapy. Methods: This study retrospectively analyzed the distribution and antibiotic resistance of pathogens based on the clinical data of BSI patients presented in the ICU of a tertiary teaching hospital from 2018 to 2020. Test performance for the prediction of pathogen species was assessed by receiver operating characteristic (ROC) analysis. Results: The analysis of the data of 382 BSI cases (10.40 cases per thousand patient day) revealed the most frequently isolated microorganisms to be Klebsiella pneumonia (11.52%), followed by Escherichia coli (9.95%), Staphylococcus epidermidis (9.95%), Candida parapsilosis (8.12%), and Enterococcus faecium (8.12%). Out of the isolated E. coli and K. pneumonia strains, 52.63, and 36.36%, respectively, were extended-spectrum ß-lactamase (ESBL) positive. The antibiotic-resistance rate of the ESBL-positive strains was 30.56% for piperacillin/tazobactam, 5.56% for imipenem, and 11.11% for tigecycline. In addition, most A. baumannii belonged to the group of multidrug-resistant (MDR) strains, with an antibiotic-resistance rate of 90.48% for meropenem and 16.00% for amikacin. However, polymyxin-resistant A. baumannii strains were not detected. Four strains of methicillin-resistant S. aureus (MRSA) (4/21, 19.05%) and one strain of vancomycin-resistant enterococci (VRE) were detected, with a resistance rate of 4.76 and 2.32%, respectively. Among the isolated 55 fungal strains, C. parapsilosis was the most common one (30/55, 56.36%), with an antibiotic-resistance rate of 5.77% for voriconazole, fluconazole, and itraconazole. The presence of amphotericin B-or flucytosine-resistant strains was not observed. Compared with the patients with Gram-positive and fungal pathogens, patients with Gram-negative bacteria exhibited the highest sequential organ failure assessment (SOFA) score (P < 0.001), lowest Glasgow Coma Scale (GCS) (P = 0.010), lowest platelet (PLT) value (P < 0.001), highest plasma creatinine (Cr) value (P = 0.016), and the highest procalcitonin (PCT) value (P < 0.001). The AUC in the ROC curve was 0.698 for the differentiation of Gram-negative BSI from Gram-positive BSI. A cutoff value of 8.47 ng/mL for PCT indicated a sensitivity of 56.9% and a specificity of 75.5%. The AUC in the ROC curve was 0.612 for the differentiation of bacteremia from fungemia. A cutoff value of 4.19 ng/mL for PCT indicated a sensitivity of 56.8% and a specificity of 62.7%. Conclusion: Among the bloodstream infection strains in ICU, Gram-negative bacteria have the highest drug resistance rate, and will cause more serious brain damage, renal function damage and thrombocytopenia. So clinician should pay more attention to the treatment of Gram-negative bacteria in patients with bloodstream infection in ICU. The test index of PCT can be used to distinguish Gram-negative bacteremia from Gram-positive and bacteremia from fungemia but not as an effective indicator, thereby indicating the need for further large-scale research.

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