RESUMEN
BACKGROUND: The prevalence of procoagulant state under prolonged hypoxic exposures and the complications and lack of specificity associated with use of existing anti-thrombotic agents have necessitated the search for safer and natural therapeutics. Codonopsis, a widely studied medicinal herb, has been reported to decrease whole blood viscosity but the bioactive ingredients involved, and their mechanism of action therein however remain to be investigated. PURPOSE: The present study aimed at evaluating the efficacy of C. clematidea root extract and mechanism of action of its bioactive constituent flavonoid, Kaempferol, in ameliorating hypobaric hypoxia induced procoagulant state. METHODS: Fingerprinting analysis of methanolic extract of C. clematidea root was performed by RP-HPLC. In vitro toxicity study was conducted using HUVEC cell line and in vivo acute and sub-acute toxicity were done according to OECD guidelines (section-4, number-420 and 407 respectively). Adult male Sprague-Dawley rats weighing 230-250â¯g were exposed to global hypoxia simulating an altitude of 7600â¯m (282â¯mmHg), in animal decompression chamber for 3, 7, 14 and 21 days for in vivo studies. Dose optimisation of the extract was done by quantification of Thromboxane A2 in the serum of hypoxic rats. C. clematidea root extract was also evaluated for its in vitro and in vivo antioxidant properties. Procoagulant changes were studied by biochemical plasma coagulation assays and expression analysis of the signalling molecules of the platelet activation cascade like vWF, platelet activation marker CD41, GpIb-IX-V (CD42), Lyn kinase, p-PI3K, p-ERK and p-PLCγ were conducted to investigate C. clematidea mediated signalling mechanisms. RESULTS: Methanolic extract of C. clematidea root showed improved antioxidant status and improvement in bleeding time and in vitro coagulation assays like pT, aPTT, INR. Decreased concentrations of D-Dimers along with that of platelet activation marker CD41 and serum concentration of Thromboxane A2 were observed in C. clematidea root extract supplemented hypoxic animals. Phosphorylation of Lyn kinase, was reduced despite increase in concentration of activating ligand vWF. CONCLUSION: C. clematidea root extract was effective in preventing hypoxia induced platelet activation and resultant procoagulant state by inhibiting Lyn kinase, a serine threonine kinase effector of vWF signalling cascade.
Asunto(s)
Codonopsis/química , Hipoxia/complicaciones , Extractos Vegetales/farmacología , Complejo GPIb-IX de Glicoproteína Plaquetaria/antagonistas & inhibidores , Familia-src Quinasas/metabolismo , Animales , Coagulación Sanguínea/efectos de los fármacos , Activación Enzimática/efectos de los fármacos , Quempferoles/farmacología , Masculino , Metanol/química , Fosforilación/efectos de los fármacos , Extractos Vegetales/química , Raíces de Plantas/química , Activación Plaquetaria/efectos de los fármacos , Complejo GPIb-IX de Glicoproteína Plaquetaria/metabolismo , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Tromboxano A2/sangreRESUMEN
A Gram-negative, facultatively anaerobic, non-motile, nonspore- forming, coccoid or rod-shaped and creamy-pigmented bacterium, designated SYP-B2100T, was isolated from the rhizospheric soil of Codonopsis clematidea in the Xinjiang Uygur Autonomous Region, China. The optimal growth occurred at 28°C, pH 5.0, in the absence of NaCl. The cells tested positive in catalase and methyl red tests but negative in oxidase, urease, gelatinase, milk coagulation, and peptonisation, H2S production, nitrate reduction, and Voges-Proskauer tests. The major isoprenoid quinone was ubiquinone-8 (Q-8). The major cellular fatty acids were C16:0 and summed feature 8. The polar lipids consisted of diphosphatidylglycerol, phosphatidylethanolamine, and phosphatidylglycerol. The 16S rRNA gene sequence of strain SYP-B2100T was the most similar to that of Rahnella inusitata DSM 30078T (96.9%) within the family Enterobacteriaceae. The genomic DNA G + C content of strain SYP-B2100T was 50.3 mol%. The combined data from the phylogenetic, morphological, physiological, biochemical, and chemotaxonomic analyses presented in this study support the conclusion that strain SYP-B2100T represents a novel species of a new genus, for which the name Edaphovirga cremea gen. nov., sp. nov. is proposed; the type strain is SYPB2100T (= CGMCC 1.5857T = DSM 105170T = KCTC 62024T).
Asunto(s)
Codonopsis/microbiología , Enterobacteriaceae/clasificación , Enterobacteriaceae/aislamiento & purificación , Técnicas de Tipificación Bacteriana , Composición de Base/genética , ADN Bacteriano/genética , Enterobacteriaceae/genética , Enterobacteriaceae/metabolismo , Ácidos Grasos/análisis , Fosfolípidos/análisis , Filogenia , ARN Ribosómico 16S/genética , Rizosfera , Microbiología del SueloRESUMEN
In an attempt to explore reasonable and impervious remedies against visceral leishmaniasis, antileishmanial potential of hydroethanolic extract of Codonopsis clematidea (HECC) and its active component, naringenin (NRG) was investigated on the basis of innocuous and immunostimulatory properties. In vitro analysis showed the ability of HECC and NRG to arrest the promastigotes in sub G0/G1 phase. Further to evaluate the protective efficacy, inbred BALB/c mice infected with L. donovani were treated with HECC and NRG for 14 days. The treated animals were sacrificed on 7th and 14th post treatment days and scrutinized for clearance of parasite, DTH response, different Th1/Th2 cytokines (IFN-γ, IL-12, TNF-α, IL-17, IL-10, IL-4), T cells and B cell responses. The expression of iNOS, NFκB and the levels of nitric oxide (NO) and reactive oxygen species was also evaluated. The toxic effect of HECC and NRG was checked in terms of biochemical parameters and histological studies. Maximum reduction in parasite load and increase in the DTH response was observed in NRG treated animals in comparison to HECC and infected control. HECC and NRG switched the host Th2 immune response to the Th1-type along with the induction of CD4+ and CD8+ T cells. CD19 B cells were found to be decreased in NRG and HECC treated animals as compared to infected control. Moreover, treatment of HECC and NRG showed no alterations in hepatic and renal enzymes which was well supported by normal architecture of liver and kidney. The mechanistic details of NRG proved that it increased the NO and ROS production by activating the NFκB and iNOS expression and thus reduced the parasite load. These findings depicted that activity of HECC might be due to the presence of NRG and that the NRG provides an encouraging alternative for the treatment of visceral leishmaniasis with the rejuvenation of immune status of the host.