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1.
Sci Rep ; 14(1): 14714, 2024 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-38926419

RESUMO

Stevia rebaudiana (stevia) is a plant in the Asteraceae that contains several biologically active compounds including the antidiabetic diterpene glycosides (e.g. stevioside, rebaudioside and dulcoside) that can serve as zero-calorie sugar alternatives. In this study, an elicitation strategy was applied using 5% polyethylene glycol (PEG), sodium chloride (NaCl; 50 and 100 mM) and gibberellic acid (2.0 and 4.0 mg/L GA3) to investigate their effect on shoot morphogenesis, and the production of phenolics, flavonoids, total soluble sugars, proline and stevioside, as well as antioxidant activity, in shoot cultures of S. rebaudiana. Herewith, the media supplemented with 2 mg/L and 4 mg/L GA3 exhibited the highest shooting response (87% and 80%). The augmentation of lower concentrations of GA3 (2 mg/L) in combination with 6-benzylaminopurine (BAP) resulted in the maximum mean shoot length (11.1 cm). The addition of 100 mM NaCl salts to the media led to the highest observed total phenolics content (TPC; 4.11 mg/g-DW compared to the control 0.52 mg/g-DW), total flavonoids content (TFC; 1.26 mg/g-DW) and polyphenolics concentration (5.39 mg/g-DW) in shoots cultured. However, the maximum antioxidant activity (81.8%) was observed in shoots raised in media treated with 50 mM NaCl. The application of 2 mg/L of GA3 resulted in the highest accumulation of proline (0.99 µg/mL) as compared to controls (0.37 µg/mL). Maximum stevioside content (71 µL/mL) was observed in cultures supplemented with 100 mM NaCl and 5% PEG, followed by the 4 mg/L GA3 treatment (70 µL/mL) as compared to control (60 µL/mL). Positive correlation was observed between GA3 and stevioside content. Notably, these two compounds are derived from a shared biochemical pathway. These results suggest that elicitation is an effective option to enhance the accumulation of steviosides and other metabolites and provides the groundwork for future industrial scale production using bioreactors.


Assuntos
Antioxidantes , Diterpenos do Tipo Caurano , Giberelinas , Glucosídeos , Brotos de Planta , Stevia , Stevia/metabolismo , Stevia/crescimento & desenvolvimento , Stevia/efeitos dos fármacos , Diterpenos do Tipo Caurano/metabolismo , Glucosídeos/metabolismo , Brotos de Planta/metabolismo , Brotos de Planta/crescimento & desenvolvimento , Brotos de Planta/efeitos dos fármacos , Giberelinas/metabolismo , Antioxidantes/metabolismo , Metabolismo Secundário , Flavonoides/metabolismo , Flavonoides/análise , Fenóis/metabolismo , Cloreto de Sódio/farmacologia , Purinas/metabolismo , Prolina/metabolismo , Polietilenoglicóis/farmacologia , Polietilenoglicóis/química , Compostos de Benzil
2.
Biochem J ; 481(12): 779-791, 2024 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-38829839

RESUMO

ent-Kaurene is a biosynthetic intermediate diterpene of phytohormone gibberellins, and is biosynthesized from geranylgeranyl diphosphate via ent-copalyl diphosphate (ent-CDP). The successive cyclization is catalyzed by two distinct diterpene synthases, ent-CDP synthase (ent-CPS) and ent-kaurene synthase (KS). Homologs of these diterpene synthase genes have been reported to be involved in the biosynthesis of specialized-metabolic diterpenoids for defense in several plant species, including rice (Oryza sativa). These diterpene synthases consist of three domains, αßγ domains. Active sites of ent-CPS exist at the interface of ß and γ domain, while those of KS are located within the α domain. We herein carried out domain-deletion experiments using several KSs and KS like enzymes (KSLs) to obtain insights into the roles of domains other than active-site domains. As previously reported in taxadiene synthase, deletion of γ or ßγ domains drastically decreased activities of specialized-metabolic OsKSL5, OsKSL8, OsKSL7 and OsKSL10 in O. sativa. However, unexpectedly, only α domains of several gibberellin-biosynthetic KSs, including OsKS1 in O. sativa, AtKS in Arabidopsis thaliana, TaKS in wheat (Triticum aestivum) and BdKS1 in Brachypodium distachyon, retained their original functions. Additionally, the specialized-metabolic OsKSL4, which is closely related to OsKS1, also functioned without its ßγ domains. Domain-swapping experiments showed that replacing ßγ domains in OsKSL7 with those from other KS/KSLs retained the OsKSL7 activity. Moreover, deletion of ßγ domains of bifunctional PpCPS/KS in moss (Physcomitrella patens) drastically impaired its KS-related activity. Thus, we demonstrate that monofunctional gibberellin-biosynthetic KSs are the unique diterpene synthases that retain their functions without ßγ domains.


Assuntos
Alquil e Aril Transferases , Giberelinas , Oryza , Proteínas de Plantas , Giberelinas/metabolismo , Alquil e Aril Transferases/metabolismo , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/química , Oryza/enzimologia , Oryza/genética , Oryza/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas de Plantas/química , Domínio Catalítico , Diterpenos do Tipo Caurano/metabolismo , Diterpenos do Tipo Caurano/química , Arabidopsis/genética , Arabidopsis/enzimologia , Arabidopsis/metabolismo , Diterpenos/metabolismo , Diterpenos/química , Domínios Proteicos , Catálise
3.
Molecules ; 29(11)2024 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-38893518

RESUMO

Oridonin (Ori) is a naturally existing diterpenoid substance that mainly exists in the Chinese medicinal plant Rabdosia rubescens. It was previously found to possess intriguing biological properties; however, the quick clearance from plasma and limited solubility in water restricts its use as a drug. Several metal-organic frameworks (MOFs), having big surfaces and large pores, have recently been considered promising drug transporters. The zeolitic imidazolate framework-8 (ZIF-8), a form of MOF consisting of 2-methylimidazole with zinc ions, is structurally stable under physiologically neutral conditions, while it can degrade at low pH values such as in tumor cells. Herein, a nanosized drug delivery system, Ori@ZIF-8, was successfully designed for encapsulating and transporting oridonin to the tumor site. The drug loading of the prepared Ori@ZIF-8 was 26.78%, and the particles' mean size was 240.5 nm. In vitro, the release of Ori@ZIF-8 exhibited acid sensitivity, with a slow release under neutral conditions and rapid release of the drug under weakly acidic conditions. According to the in vitro anti-tumor experiments, Ori@ZIF-8 produced higher cytotoxicity than free Ori and induced apoptosis in A549 cancer cells. In conclusion, Ori@ZIF-8 could be a potential pH-responsive carrier to accurately release more oridonins at the tumor site.


Assuntos
Diterpenos do Tipo Caurano , Estruturas Metalorgânicas , Diterpenos do Tipo Caurano/química , Diterpenos do Tipo Caurano/farmacologia , Estruturas Metalorgânicas/química , Humanos , Concentração de Íons de Hidrogênio , Sistemas de Liberação de Medicamentos , Liberação Controlada de Fármacos , Portadores de Fármacos/química , Células A549 , Linhagem Celular Tumoral , Zeolitas/química , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Antineoplásicos/farmacologia , Antineoplásicos/química , Sobrevivência Celular/efeitos dos fármacos , Imidazóis
4.
Ren Fail ; 46(1): 2347462, 2024 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-38832497

RESUMO

Diabetic nephropathy (DN) is one of the most serious and frequent complications among diabetes patients and presently constitutes vast the cases of end-stage renal disease worldwide. Tubulointerstitial fibrosis is a crucial factor related to the occurrence and progression of DN. Oridonin (Ori) is a diterpenoid derived from rubescens that has diverse pharmacological properties. Our previous study showed that Ori can protect against DN by decreasing the inflammatory response. However, whether Ori can alleviate renal fibrosis in DN remains unknown. Here, we investigated the mechanism through which Ori affects the Wnt/ß-catenin signaling pathway in diabetic rats and human proximal tubular epithelial cells (HK-2) exposed to high glucose (HG) levels. Our results revealed that Ori treatment markedly decreased urinary protein excretion levels, improved renal function and alleviated renal fibrosis in diabetic rats. In vitro, HG treatment increased the migration of HK-2 cells while reducing their viability and proliferation rate, and treatment with Ori reversed these changes. Additionally, the knockdown of ß-catenin arrested cell migration and reduced the expression levels of Wnt/ß-catenin signaling-related molecules (Wnt4, p-GSK3ß and ß-catenin) and fibrosis-related molecules (α-smooth muscle actin, collagen I and fibronectin), and Ori treatment exerted an effect similar to that observed after the knockdown of ß-catenin. Furthermore, the combination of Ori treatment and ß-catenin downregulation exerted more pronounced biological effects than treatment alone. These findings may provide the first line of evidence showing that Ori alleviates fibrosis in DN by inhibiting the Wnt/ß-catenin signaling pathway and thereby reveal a novel therapeutic avenue for treating tubulointerstitial fibrosis.


Assuntos
Diabetes Mellitus Experimental , Nefropatias Diabéticas , Diterpenos do Tipo Caurano , Fibrose , Via de Sinalização Wnt , Animais , Humanos , Masculino , Ratos , beta Catenina/metabolismo , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/tratamento farmacológico , Nefropatias Diabéticas/tratamento farmacológico , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/etiologia , Diterpenos do Tipo Caurano/farmacologia , Diterpenos do Tipo Caurano/uso terapêutico , Fibrose/tratamento farmacológico , Rim/patologia , Rim/efeitos dos fármacos , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/patologia , Túbulos Renais Proximais/metabolismo , Ratos Sprague-Dawley , Via de Sinalização Wnt/efeitos dos fármacos
5.
Int J Radiat Biol ; 100(7): 1104-1115, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38870412

RESUMO

PURPOSE: Stevia rebaudiana Bertoni is a perennial herb, widely used as a natural sweetener around the globe. The key compounds responsible for its sweetness includes stevioside and rebaudioside-A. In order to improve these steviol glycosides, the present study was initiated to study the effect of induced mutagenesis on growth parameters, steviol glycosides and nuclear DNA content in Stevia rebaudiana Bertoni using ten doses of gamma-rays (5-100 kR). MATERIALS AND METHODS: Healthy seeds of 'Madhuguna' variety of Stevia rebaudiana Bertoni developed and maintained at stevia breeding farm, Agrotechnology division, CSIR-Institute of Himalayan Bioresource Technology, Palampur (HP), India were irradiated with ten doses of gamma rays (600 seeds each/dose) ranging from 5 kR to 100 kR (i.e., 5, 10, 15, 20, 30, 40, 50, 60, 80 and 100 kR) using Co60 gamma irradiation chamber at CCS Haryana Agricultural University, Hisar, (Haryana), India. RESULTS: Significant variations were recorded for all the seedling traits studied while major impact was noticed on the seedling after reaching the cotyledonary stage and doses above 40 kR showed absolute mortality of the seedlings. Based on probit analysis, the optimum LD50 dose lies in the range of 20-23 kR. Glycosidic profiling of 296 mutants using high-performance liquid chromatography showed decreased total steviol glycoside content with increased radiation dose. Doses 5 kR and 10 kR, were found to be effective in increasing the overall glycosidic content. A total of 72 promising mutants were also screened for increased rebaudioside-A stevioside ratio. Comparison of nuclear DNA content using flow cytometry revealed a similar decrease in the total nuclear DNA content with increase in dosage of gamma rays. The average genome size at 5, 10, 15, 20 and 30 kR treatments were 2.72, 2.69, 2.68, 2.70 and 2.66 pg as compared to 2.72 pg in control. CONCLUSIONS: Mild dose of gamma rays (5 and 10 kR) in stevia were found to be effective in improving the mean steviol glycoside content and may be used in future stevia mutation programmes.


Assuntos
Diterpenos do Tipo Caurano , Raios gama , Stevia , Stevia/efeitos da radiação , Tolerância a Radiação , Glucosídeos , Relação Dose-Resposta à Radiação
6.
Biomed Chromatogr ; 38(8): e5943, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38890009

RESUMO

In this study, a targeted nanocarrier was developed by functionalizing graphene oxide with polyethyleneimine and folic acid, intended for loading oridonin. The nanocarrier was successfully synthesized and characterized using an ultraviolet spectrum, Fourier transform infrared spectroscopy and scanning electron microscopy. The nanocarrier demonstrated a remarkable oridonin loading capacity, reaching 424.8 µg/mg, as determined by ultra-high performance liquid chromatography. In vitro drug release experiments exhibited a pH-dependent release profile, with a higher cumulative release in an acidic environment. The release mechanism followed the Ritger-Peppas equation model. Cytotoxicity assays indicated minimal toxicity of the nanocarrier. Enhanced cellular uptake by MCF7 cells was observed for carriers functionalized with folate and polyethyleneimine. These findings highlight the potential of functionalized graphene oxide as a promising carrier for oridonin delivery in biomedical applications.


Assuntos
Neoplasias da Mama , Diterpenos do Tipo Caurano , Portadores de Fármacos , Grafite , Grafite/química , Diterpenos do Tipo Caurano/química , Diterpenos do Tipo Caurano/farmacologia , Humanos , Células MCF-7 , Portadores de Fármacos/química , Neoplasias da Mama/tratamento farmacológico , Feminino , Antineoplásicos/farmacologia , Antineoplásicos/química , Liberação Controlada de Fármacos , Sobrevivência Celular/efeitos dos fármacos , Ácido Fólico/química , Nanopartículas/química , Cromatografia Líquida de Alta Pressão/métodos
7.
Cell Mol Biol (Noisy-le-grand) ; 70(6): 108-113, 2024 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-38836673

RESUMO

Oridonin belongs to a small molecule from the Chinese herb Rabdosia rubescens with potent anticancer activity. In spite of the lncRNA AFAP1-AS1 has been proven to exert promoting function in lung cancer, its relationship with oridonin in lung cancer is obscure. Therefore, our study planned to explore the potential of oridonin in lung cancer as well as unveil the regulatory mechanism of oridonin on AFAP1-AS1 in lung cancer cells. In the present study, oridonin inhibited lung cancer cell proliferation, migration, as well as invasion, as evidenced by MTT, wound healing, as well as transwell assays. Besides, we observed that oridonin could downregulate AFAP1-AS1 expression, and overexpressed AFAP1-AS1 could reverse the repressive effects of oridonin on lung cancer cell proliferation, migration, as well as invasion. More importantly, we found that AFAP1-AS1 could bind to IGF2BP1 through starBase prediction and RIP assay. The expression level of IGF2BP1 was also reduced by oridonin treatment but reversed after AFAP1-AS1 overexpression. Additionally, we proved that overexpressed IGF2BP1 could reverse the repressive impacts of oridonin on lung cancer cell proliferation, migration, as well as invasion. Further, in vivo experiments validated the repressive role of oridonin on tumor growth of lung cancer. Together, oridonin inhibits lung cancer cell proliferation as well as migration by modulating AFAP1-AS1/IGF2BP1, and AFAP1-AS1/IGF2BP1 possesses the potential to be a promising therapy targeting for lung cancer, especially in oridonin treatment.


Assuntos
Movimento Celular , Proliferação de Células , Diterpenos do Tipo Caurano , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares , RNA Longo não Codificante , Proteínas de Ligação a RNA , Diterpenos do Tipo Caurano/farmacologia , Humanos , Proliferação de Células/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Animais , Camundongos Nus , Camundongos , Camundongos Endogâmicos BALB C , Invasividade Neoplásica , Células A549
8.
Food Chem ; 453: 139622, 2024 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-38761729

RESUMO

For health and safety reasons, the search for green, healthy, and low-calorie sweeteners with good taste has become the demand of many consumers. Furthermore, the need for sugar substitutes of natural origin has increased dramatically. In this review, we briefly discussed the safety and health benefits of stevia sweeteners and enumerated some examples of physiological functions of steviol glycosides (SGs), such as anti-inflammatory, anti-obesity, antihypertensive, anti-diabetes, and anticaries, citing various evidence related to their application in the food industry. The latest advances in emerging technologies for extracting and purifying SGs and the process variables and operational strategies were discussed. The impact of the extraction methods and their comparison against the conventional techniques have also been demonstrated. These technologies use minimal energy solvents and simplify subsequent purification stages, making viable alternatives suitable for a possible industrial application. Furthermore, we also elucidated the potential for advancing and applying the natural sweeteners SGs.


Assuntos
Diterpenos do Tipo Caurano , Extratos Vegetais , Stevia , Edulcorantes , Stevia/química , Diterpenos do Tipo Caurano/isolamento & purificação , Diterpenos do Tipo Caurano/química , Edulcorantes/isolamento & purificação , Edulcorantes/química , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Humanos , Glucosídeos/isolamento & purificação , Glucosídeos/química , Animais , Glicosídeos/isolamento & purificação , Glicosídeos/química
9.
Cell Immunol ; 401-402: 104838, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38810591

RESUMO

BACKGROUND: The NOD-like receptor protein 3 (NLRP3) mediated pyroptosis of macrophages is closely associated with liver ischemia reperfusion injury (IRI). As a covalent inhibitor of NLRP3, Oridonin (Ori), has strong anti-inflammasome effect, but its effect and mechanisms for liver IRI are still unknown. METHODS: Mice and liver macrophages were treated with Ori, respectively. Co-IP and LC-MS/MS analysis of the interaction between PKM2 and NLRP3 in macrophages. Liver damage was detected using H&E staining. Pyroptosis was detected by WB, TEM, and ELISA. RESULTS: Ori ameliorated liver macrophage pyroptosis and liver IRI. Mechanistically, Ori inhibited the interaction between pyruvate kinase M2 isoform (PKM2) and NLRP3 in hypoxia/reoxygenation(H/R)-induced macrophages, while the inhibition of PKM2/NLRP3 reduced liver macrophage pyroptosis and liver IRI. CONCLUSION: Ori exerted protective effects on liver IRI via suppressing PKM2/NLRP3-mediated liver macrophage pyroptosis, which might become a potential therapeutic target in the clinic.


Assuntos
Diterpenos do Tipo Caurano , Fígado , Macrófagos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR , Piroptose , Traumatismo por Reperfusão , Animais , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Piroptose/efeitos dos fármacos , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/tratamento farmacológico , Camundongos , Fígado/metabolismo , Fígado/efeitos dos fármacos , Fígado/patologia , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Diterpenos do Tipo Caurano/farmacologia , Masculino , Piruvato Quinase/metabolismo , Inflamassomos/metabolismo , Inflamassomos/efeitos dos fármacos , Hepatopatias/metabolismo , Hepatopatias/tratamento farmacológico
10.
Biomed Pharmacother ; 175: 116684, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38713951

RESUMO

Chinese herbs have been used to treat small-cell lung cancer (SCLC) due to their low toxicity and significant efficacy. This study focused on oridonin, a natural compound extracted from Rabdosia rubescens, and aimed to investigate its potential antitumor activity on SCLC and to evaluate the synergistic effect of combining oridonin with other small molecules. In this study, oridonin exhibited a dual effect. At lower concentrations, it suppressed the cell viability of SCLC cells (H1688 and H446). At high concentrations, oridonin induced SCLC cell apoptosis, damaged HBE cells in vitro and compromised the function of the liver and heart in vivo. The lower concentration of oridonin induced autophagy by enhancing the expression of p62 and the LC3B-II/LC3B-I ratio. This phenomenon might be associated with the activation of the protein kinase RNA-like ER kinase (PERK)/eukaryotic initiation factor 2 alpha (eIF2α)/growth arrest and DNA damage-inducible gene 153 (CHOP/GAD153) pathway. Therefore, the combined effect of oridonin with GSK2606414 or 3- methyladenine increased apoptosis in SCLC cells and reduced tumor growth. A similar phenomenon was observed after oridonin was combined with p62 or CHOP RNA interference treatment. Simultaneously, the combination of oridonin and GSK2606414 exhibited therapeutic efficacy without manifesting adverse effects. Our findings suggest that oridonin at lower concentrations can induce autophagy by activating the PERK/eIF2α/CHOP signaling pathway. The inhibition of the PERK/eIF2α/CHOP pathway could enhance oridonin therapeutic responses by triggering apoptosis. The novel therapeutic approach of combining oridonin with a PERK inhibitor is promising as a strategy for the treatment of SCLC.


Assuntos
Apoptose , Autofagia , Diterpenos do Tipo Caurano , Fator de Iniciação 2 em Eucariotos , Neoplasias Pulmonares , Transdução de Sinais , Carcinoma de Pequenas Células do Pulmão , Fator de Transcrição CHOP , eIF-2 Quinase , Diterpenos do Tipo Caurano/farmacologia , Autofagia/efeitos dos fármacos , Fator de Transcrição CHOP/metabolismo , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Carcinoma de Pequenas Células do Pulmão/patologia , Carcinoma de Pequenas Células do Pulmão/metabolismo , eIF-2 Quinase/metabolismo , Apoptose/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Linhagem Celular Tumoral , Fator de Iniciação 2 em Eucariotos/metabolismo , Animais , Transdução de Sinais/efeitos dos fármacos , Camundongos Nus , Camundongos Endogâmicos BALB C , Camundongos , Ensaios Antitumorais Modelo de Xenoenxerto , Sobrevivência Celular/efeitos dos fármacos , Sinergismo Farmacológico , Masculino
11.
Int Immunopharmacol ; 134: 112247, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38759374

RESUMO

BACKGROUND: Epilepsy is a chronic disabling disease poorly controlled by available antiseizure medications. Oridonin, a bioactive alkaloid with anti-inflammatory properties and neuroprotective effects, can inhibit the increased excitability of neurons caused by glutamate accumulation at the cellular level. However, whether oridonin affects neuronal excitability and whether it has antiepileptic potential has not been reported in animal models or clinical studies. METHOD: Pentylenetetrazol was injected into mice to create a model of chronic epilepsy. Seizure severity was assessed using the Racine scale, and the duration and latency of seizures were observed. Abnormal neuronal discharge was detected using electroencephalography, and neuronal excitability was assessed using calcium imaging. Damage to hippocampal neurons was evaluated using Hematoxylin-Eosin and Nissl staining. The expression of the NOD-like receptor thermal protein domain associated protein 3 (NLRP3) inflammasome and other pyroptosis-related proteins was determined using western blotting and immunofluorescence. A neuronal pyroptosis model was established using the supernatant of BV2 cells treated with lipopolysaccharide and adenosine triphosphate to stimulate hippocampal neurons. RESULTS: Oridonin (1 and 5 mg/kg) reduced neuronal damage, increased the latency of seizures, and shortened the duration of fully kindled seizures in chronic epilepsy model mice. Oridonin decreased abnormal discharge during epileptic episodes and suppressed increased neuronal excitability. In vitro experiments showed that oridonin alleviated pyroptosis in hippocampal HT22 neurons. CONCLUSION: Oridonin exerts neuroprotective effects by inhibiting pyroptosis through the NLRP3/caspase-1 pathway in chronic epilepsy model mice. It also reduces pyroptosis in hippocampal neurons in vitro, suggesting its potential as a therapy for epilepsy.


Assuntos
Anticonvulsivantes , Modelos Animais de Doenças , Diterpenos do Tipo Caurano , Epilepsia , Hipocampo , Proteína 3 que Contém Domínio de Pirina da Família NLR , Neurônios , Fármacos Neuroprotetores , Piroptose , Animais , Diterpenos do Tipo Caurano/farmacologia , Diterpenos do Tipo Caurano/uso terapêutico , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Epilepsia/tratamento farmacológico , Piroptose/efeitos dos fármacos , Camundongos , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Masculino , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Hipocampo/patologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Neurônios/metabolismo , Pentilenotetrazol , Camundongos Endogâmicos C57BL , Inflamassomos/metabolismo , Inflamassomos/efeitos dos fármacos , Linhagem Celular , Convulsões/tratamento farmacológico
12.
Eur J Pharmacol ; 975: 176656, 2024 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-38754536

RESUMO

Cancer stem cells (CSCs) drive malignant tumor progression, recurrence, and metastasis with unique characteristics, including self-renewal and resistance to conventional treatments. Conventional differentiation inducers, although promising, have limited cytotoxicity and may inadvertently enhance CSC stemness. To address these challenges, ongoing efforts are dedicated to developing strategies that can effectively combine both cytotoxicity and differentiation-inducing effects. In this study, we introduce oridonin (Ori), a small molecule with dual differentiation-inducing and cytotoxicity properties capable of eliminating tumor CSCs. We isolated CSCs in B16F10 cells using the Hoechst side population method and assessed the differentiation effect of Ori. Ori's differentiation-inducing effect was further evaluated using human acute promyelocytic leukemia. The cytotoxic potential of Ori against MCF-7 and B16F10 cell lines was assessed through various methods. In vivo anti-tumor and anti-CSC efficacy of Ori was investigated using mouse melanoma and CSCs melanoma models. Safety evaluation included zebrafish embryotoxicity and mouse acute toxicity experiments. As a result, Ori effectively dismantles tumorspheres, inhibits proliferation, and reduces the expression of CSC-specific markers. It induces significant differentiation, especially in the case of NB4. Additionally, Ori upregulates TP53 expression, mitigates the hypoxic tumor microenvironment, suppresses stemness, and inhibits PD-L1 expression, prompting a robust anti-cancer immune response. Ori demonstrates pronounced cytotoxicity, inducing notable pro-apoptotic effects on B16F10 and MCF-7 cells, with specific triggering of mitochondrial apoptosis. Importantly, Ori maintains a commendable biosafety record. The dual-action prowess of Ori not only induces the differentiation of CSCs but also dispatches differentiated and residual tumor cells, effectively thwarting the relentless march of tumor progression.


Assuntos
Diferenciação Celular , Diterpenos do Tipo Caurano , Células-Tronco Neoplásicas , Peixe-Zebra , Diterpenos do Tipo Caurano/farmacologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Animais , Humanos , Diferenciação Celular/efeitos dos fármacos , Camundongos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Antineoplásicos/farmacologia , Proteína Supressora de Tumor p53/metabolismo , Células MCF-7 , Melanoma Experimental/patologia , Melanoma Experimental/tratamento farmacológico , Leucemia Promielocítica Aguda/patologia , Leucemia Promielocítica Aguda/tratamento farmacológico , Feminino
13.
J Nanobiotechnology ; 22(1): 299, 2024 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-38812031

RESUMO

BACKGROUND: Discrepancies in the utilization of reactive oxygen species (ROS) between cancer cells and their normal counterparts constitute a pivotal juncture for the precise treatment of cancer, delineating a noteworthy trajectory in the field of targeted therapies. This phenomenon is particularly conspicuous in the domain of nano-drug precision treatment. Despite substantial strides in employing nanoparticles to disrupt ROS for cancer therapy, current strategies continue to grapple with challenges pertaining to efficacy and specificity. One of the primary hurdles lies in the elevated levels of intracellular glutathione (GSH). Presently, predominant methods to mitigate intracellular GSH involve inhibiting its synthesis or promoting GSH efflux. However, a conspicuous gap remains in the absence of a strategy capable of directly and efficiently clearing GSH. METHODS: We initially elucidated the chemical mechanism underpinning oridonin, a diminutive pharmacological agent demonstrated to perturb reactive oxygen species, through its covalent interaction with glutathione. Subsequently, we employed the incorporation of maleimide-liposomes, renowned for their capacity to disrupt the ROS delivery system, to ameliorate the drug's water solubility and pharmacokinetics, thereby enhancing its ROS-disruptive efficacy. In a pursuit to further refine the targeting for acute myeloid leukemia (AML), we harnessed the maleic imide and thiol reaction mechanism, facilitating the coupling of Toll-like receptor 2 (TLR2) peptides to the liposomes' surface via maleic imide. This strategic approach offers a novel method for the precise removal of GSH, and its enhancement endeavors are directed towards fortifying the precision and efficacy of the drug's impact on AML targets. RESULTS: We demonstrated that this peptide-liposome-small molecule machinery targets AML and consequently induces cell apoptosis both in vitro and in vivo through three disparate mechanisms: (I) Oridonin, as a Michael acceptor molecule, inhibits GSH function through covalent bonding, triggering an initial imbalance of oxidative stress. (II) Maleimide further induces GSH exhaustion, aggravating redox imbalance as a complementary augment with oridonin. (III) Peptide targets TLR2, enhances the directivity and enrichment of oridonin within AML cells. CONCLUSION: The rationally designed nanocomplex provides a ROS drug enhancement and targeted delivery platform, representing a potential solution by disrupting redox balance for AML therapy.


Assuntos
Diterpenos do Tipo Caurano , Glutationa , Leucemia Mieloide Aguda , Lipossomos , Espécies Reativas de Oxigênio , Diterpenos do Tipo Caurano/química , Diterpenos do Tipo Caurano/farmacologia , Glutationa/metabolismo , Glutationa/química , Lipossomos/química , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Humanos , Espécies Reativas de Oxigênio/metabolismo , Animais , Camundongos , Linhagem Celular Tumoral , Receptor 2 Toll-Like/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/química , Apoptose/efeitos dos fármacos
14.
J Med Chem ; 67(11): 9406-9430, 2024 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-38751194

RESUMO

Targeting NLRP3 inflammasome with inhibitors is a novel strategy for NLRP3-driven diseases. Herein, hit compound 5 possessing an attractive skeleton was identified from our in-house database of oridonin, and then a potential lead compound 32 was obtained by optimization of 5, displaying two-digit nanomolar inhibition on NLRP3. Moreover, compound 32 showed enhanced safety index (SI) relative to oridonin (IC50 = 77.2 vs 780.4 nM, SI = 40.5 vs 8.5) and functioned through blocking ASC oligomerization and interaction of NLRP3-ASC/NEK7, thereby suppressing NLRP3 inflammasome assembly and activation. Furthermore, diverse agonists-induced activations of NLRP3 could be impeded by compound 32 without altering NLRC4 or AIM2 inflammasome. Crucially, compound 32 possessed tolerable pharmaceutical properties and significant anti-inflammatory activity in MSU-induced gouty arthritis model. Therefore, this work enriched the SAR of NLRP3 inflammasome inhibitors and provided a potential candidate for the treatment of NLRP3-associated diseases.


Assuntos
Anti-Inflamatórios , Diterpenos do Tipo Caurano , Inflamassomos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Diterpenos do Tipo Caurano/farmacologia , Diterpenos do Tipo Caurano/química , Diterpenos do Tipo Caurano/uso terapêutico , Diterpenos do Tipo Caurano/síntese química , Inflamassomos/metabolismo , Inflamassomos/antagonistas & inibidores , Animais , Humanos , Camundongos , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/química , Anti-Inflamatórios/uso terapêutico , Anti-Inflamatórios/síntese química , Relação Estrutura-Atividade , Masculino , Produtos Biológicos/farmacologia , Produtos Biológicos/química , Produtos Biológicos/síntese química , Camundongos Endogâmicos C57BL , Quinases Relacionadas a NIMA/antagonistas & inibidores , Quinases Relacionadas a NIMA/metabolismo
15.
Zhongguo Zhong Yao Za Zhi ; 49(9): 2410-2421, 2024 May.
Artigo em Chinês | MEDLINE | ID: mdl-38812142

RESUMO

Sequential catalysis by ent-copalyl diphosphate(CPS) and ent-kaurene synthase(KS) is a critical step for plants to initiate the biosynthesis of gibberellin with geranylgeranyl pyrophosphate(GGPP) as the substrate. This study mined the transcriptome data of Stellera chamaejasme and cloned two key diterpene synthase genes, SchCPS and SchKS, involved in the gibberellin pathway. The two genes had the complete open reading frames of 2 595 bp and 1 701 bp, encoding two hydrophilic proteins composed of 864 and 566 amino acid residues and with the relative molecular mass of 97.9 kDa and 64.6 kDa and the theoretical isoelectric points of 5.61 and 6.12, respectively. Sequence comparison and phylogenetic tree showed that SchCPS contained LHS, PNV, and DxDD motifs conserved in the CPS family and was categorized in the TPS-c subfamily, while SchKS contained DDxxD, NSE/DTE and PIx motifs conserved in the KS family and was categorized in the TPS-e subfamily. Functional validation showed that SchCPS catalyzed the protonation and cyclization of GGPP to ent-CPP, while SchKS acted on ent-CPP dephosphorylation and re-cyclization to ent-kaurene. In this study, the full-length sequences of SchCPS and SchKS were cloned and functionally verified for the first time, which not only enriched the existing CPS and KS gene libraries but also laid a foundation for the cloning and biosynthesis pathway analysis of more genes involved in the synthesis of active components in S. chamaejasme.


Assuntos
Alquil e Aril Transferases , Filogenia , Proteínas de Plantas , Thymelaeaceae , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Alquil e Aril Transferases/química , Thymelaeaceae/genética , Thymelaeaceae/enzimologia , Thymelaeaceae/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas de Plantas/química , Sequência de Aminoácidos , Diterpenos do Tipo Caurano/metabolismo , Diterpenos do Tipo Caurano/química , Alinhamento de Sequência , Clonagem Molecular
16.
Endocrinol Diabetes Metab ; 7(3): e00482, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38556697

RESUMO

BACKGROUND: Stevioside (SV) with minimal calories is widely used as a natural sweetener in beverages due to its high sweetness and safety. However, the effects of SV on glucose uptake and the pyruvate dehydrogenase kinase isoenzyme (PDK4) as an important protein in the regulation of glucose metabolism, remain largely unexplored. In this study, we used C2C12 skeletal muscle cells that was induced by palmitic acid (PA) to assess the effects and mechanisms of SV on glucose uptake and PDK4. METHODS: The glucose uptake of C2C12 cells was determined by 2-NBDG; expression of the Pdk4 gene was measured by quantitative real-time PCR; and expression of the proteins PDK4, p-AMPK, TBC1D1 and GLUT4 was assessed by Western blotting. RESULTS: In PA-induced C2C12 myotubes, SV could significantly promote cellular glucose uptake by decreasing PDK4 levels and increasing p-AMPK and TBC1D1 levels. SV could promote the translocation of GLUT4 from the cytoplasm to the cell membrane in cells. Moreover, in Pdk4-overexpressing C2C12 myotubes, SV decreased the level of PDK4 and increased the levels of p-AMPK and TBC1D1. CONCLUSION: SV was found to ameliorate PA-induced abnormal glucose uptake via the PDK4/AMPK/TBC1D1 pathway in C2C12 myotubes. Although these results warranted further investigation for validation, they may provide some evidence of SV as a safe natural sweetener for its use in sugar-free beverages to prevent and control T2DM.


Assuntos
Proteínas Quinases Ativadas por AMP , Diterpenos do Tipo Caurano , Glucosídeos , Ácido Palmítico , Ácido Palmítico/metabolismo , Ácido Palmítico/farmacologia , Piruvato Desidrogenase Quinase de Transferência de Acetil/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Proteínas Quinases Ativadas por AMP/farmacologia , Músculo Esquelético/metabolismo , Glucose/metabolismo , Glucose/farmacologia , Fibras Musculares Esqueléticas/metabolismo , Edulcorantes/farmacologia , Edulcorantes/metabolismo
17.
J Agric Food Chem ; 72(14): 8140-8148, 2024 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-38563232

RESUMO

Rebaudioside (Reb) M is an important sweetener with high sweetness, but its low content in Stevia rebaudiana and low catalytic capacity of the glycosyltransferases in heterologous microorganisms limit its production. In order to improve the catalytic efficiency of the conversion of stevioside to Reb M by Saccharomyces cerevisiae, several key issues must be resolved including knocking out endogenous hydrolases, enhancing glycosylation, and extending the enzyme catalytic process. Herein, endogenous glycosyl hydrolase SCW2 was knocked out in S. cerevisiae. The glycosylation process was enhanced by screening glycosyltransferases, and UGT91D2 from S. rebaudiana was identified as the optimum glycosyltransferase. The UDP-glucose supply was enhanced by overexpressing UGP1, and co-expressing UGT91D2 and UGT76G1 achieved efficient conversion of stevioside to Reb M. In order to extend the catalytic process, the silencing information regulator 2 (SIR2) which can prolong the growth cycle of S. cerevisiae was introduced. Finally, combining these modifications produced 12.5 g/L Reb M and the yield reached 77.9% in a 5 L bioreactor with 10.0 g/L stevioside, the highest titer from steviol glycosides to Reb M reported to date. The engineered strain could facilitate the industrial production of Reb M, and the strategies provide references for the production of steviol glycosides.


Assuntos
Diterpenos do Tipo Caurano , Stevia , Trissacarídeos , Saccharomyces cerevisiae/genética , Difosfato de Uridina , Hidrolases , Glucosídeos , Glicosiltransferases/genética , Glicosídeos , Folhas de Planta
18.
Int J Mol Sci ; 25(7)2024 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-38612735

RESUMO

The antitumor activity of different ent-kaurane diterpenes has been extensively studied. Several investigations have demonstrated the excellent antitumor activity of synthetic derivatives of the diterpene atractyligenin. In this research, a series of new synthetic amides and their 15,19-di-oxo analogues obtained from atractyligenin by modifying the C-2, C-15, and C-19 positions were designed in order to dispose of a set of derivatives with different substitutions at the amidic nitrogen. Using different concentrations of the obtained compounds (10-300 µM) a reduction in cell viability of HCT116 colon cancer cells was observed at 48 h of treatment. All the di-oxidized compounds were more effective than their alcoholic precursors. The di-oxidized compounds had already reduced the viability of two colon cancer cells (HCT116 and Caco-2) at 24 h when used at low doses (2.5-15 µM), while they turned out to be poorly effective in differentiated Caco-2 cells, a model of polarized enterocytes. The data reported here provide evidence that di-oxidized compounds induced apoptotic cell death, as demonstrated by the appearance of condensed and fragmented DNA in treated cells, as well as the activation of caspase-3 and fragmentation of its target PARP-1.


Assuntos
Atractilosídeo/análogos & derivados , Neoplasias do Colo , Diterpenos do Tipo Caurano , Humanos , Diterpenos do Tipo Caurano/farmacologia , Células CACO-2 , Neoplasias do Colo/tratamento farmacológico , Amidas , Apoptose
19.
J Med Chem ; 67(8): 6749-6768, 2024 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-38572607

RESUMO

Cardiovascular diseases (CVDs) persist as the predominant cause of mortality, urging the exploration of innovative pharmaceuticals. Mitochondrial dysfunction stands as a pivotal contributor to CVDs development. Sirtuin 3 (SIRT3), a prominent mitochondrial deacetylase known for its crucial role in protecting mitochondria against damage and dysfunction, has emerged as a promising therapeutic target for CVDs treatment. Utilizing isosteviol, a natural ent-beyerene diterpenoid, 24 derivatives were synthesized and evaluated in vivo using a zebrafish model, establishing a deduced structure-activity relationship. Among these, derivative 5v exhibited significant efficacy in doxorubicin-induced cardiomyopathy in zebrafish and murine models. Subsequent investigations revealed that 5v selectively elevated SIRT3 expression, leading to the upregulation of SOD2 and OPA1 expression, effectively preventing mitochondrial dysfunction, mitigating oxidative stress, and preserving cardiomyocyte viability. As a novel structural class of SIRT3 activators with robust therapeutic effects, 5v emerges as a promising candidate for further drug development.


Assuntos
Cardiotônicos , Diterpenos do Tipo Caurano , Desenho de Fármacos , Sirtuína 3 , Peixe-Zebra , Animais , Sirtuína 3/metabolismo , Sirtuína 3/antagonistas & inibidores , Diterpenos do Tipo Caurano/farmacologia , Diterpenos do Tipo Caurano/síntese química , Diterpenos do Tipo Caurano/química , Diterpenos do Tipo Caurano/uso terapêutico , Cardiotônicos/farmacologia , Cardiotônicos/síntese química , Cardiotônicos/química , Cardiotônicos/uso terapêutico , Relação Estrutura-Atividade , Camundongos , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Humanos , Estresse Oxidativo/efeitos dos fármacos , Doxorrubicina/farmacologia
20.
Molecules ; 29(8)2024 Apr 19.
Artigo em Inglês | MEDLINE | ID: mdl-38675686

RESUMO

Stevia rebaudiana Bertoni is a plant native to South America that has gathered much interest in recent decades thanks to diterpene glycosides, called steviosides, which it produces. These compounds are characterised by their sweetness, which is 250-300 times higher than saccharose, and they contain almost no caloric value. Stevia is currently also grown outside the South American continent, in various countries characterised by warm weather. This research aimed to determine whether it is viable to grow Stevia rebaudiana plants in Poland, a country characterised by a cooler climate than the native regions for stevia plants. Additionally, the impact of adding various dosages and forms of nitrogen fertiliser was analysed. It was determined that Stevia rebaudiana grown in Poland is characterised by a rather low concentration of steviosides, although proper nitrogen fertilisation can improve various characteristics of the grown plants. The addition of 100 kg or 150 kg of nitrogen per hectare of the field in the form of urea or ammonium nitrate increased the yield of the stevia plants. The stevioside content can be increased by applying fertilisation using 100 kg or 150 kg of nitrogen per hectare in the form of ammonium sulfate. The total yield of the stevia plants grown in Poland was lower than the yield typically recorded in warmer countries, and the low concentration of steviosides in the plant suggests that more research about growing Stevia rebaudiana in Poland would be needed to develop profitable methods of stevia cultivation.


Assuntos
Fertilizantes , Nitrogênio , Stevia , Stevia/química , Stevia/crescimento & desenvolvimento , Polônia , Nitrogênio/análise , Fertilizantes/análise , Diterpenos do Tipo Caurano/análise , Diterpenos do Tipo Caurano/química , Glucosídeos/análise , Glucosídeos/química , Nitratos/análise , Nitratos/química
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