Platelet rich fibrin and simvastatin-loaded pectin-based 3D printed-electrospun bilayer scaffold for skin tissue regeneration.

Designing multifunctional wound dressings is a prerequisite to prevent infection and stimulate healing. In this study, a bilayer scaffold (BS) with a top layer (TL) comprising 3D printed pectin/polyacrylic acid/platelet rich fibrin hydrogel (Pec/PAA/PRF) and a bottom nanofibrous layer (NL) containing Pec/PAA/simvastatin (SIM) was produced. The biodegradable and biocompatible polymers Pec and PAA were cross-linked to form hydrogels via Ca2+ activation through galacturonate linkage and chelation, respectively. PRF as an autologous growth factor (GF) source and SIM together augmented angiogenesis and neovascularization. Because of 3D printing, the BS possessed a uniform distribution of PRF in TL and an average fiber diameter of 96.71 ± 18.14 nm was obtained in NL. The Young's modulus of BS was recorded as 6.02 ± 0.31 MPa and its elongation at break was measured as 30.16 ± 2.70 %. The wound dressing gradually released growth factors over 7 days of investigation. Furthermore, the BS significantly outperformed other groups in increasing cell viability and in vivo wound closure rate (95.80 ± 3.47 % after 14 days). Wounds covered with BS healed faster with more collagen deposition and re-epithelialization. The results demonstrate that the BS can be a potential remedy for skin tissue regeneration.

[Diosgenin alleviates NAFLD induced by a high-fat diet in rats via mTOR/SREBP-1c/HSP60/MCAD/SCAD signaling pathway].

This study aims to observe the effects of diosgenin on the expression of mammalian target of rapamycin(mTOR), sterol regulatory element-binding protein-1c(SREBP-1c), heat shock protein 60(HSP60), medium-chain acyl-CoA dehydrogenase(MCAD), and short-chain acyl-CoA dehydrogenase(SCAD) in the liver tissue of the rat model of non-alcoholic fatty liver disease(NAFLD) and explore the mechanism of diosgenin in alleviating NAFLD. Forty male SD rats were randomized into five groups: a control group, a model group, low-(150 mg·kg~(-1)·d~(-1)) and high-dose(300 mg·kg~(-1)·d~(-1)) diosgenin groups, and a simvastatin(4 mg·kg~(-1)·d~(-1)) group. The rats in the control group were fed with a normal diet, while those in the other four groups were fed with a high-fat diet. After feeding for 8 weeks, the body weight of rats in the high-fat diet groups increased significantly. After that, the rats were administrated with the corresponding dose of diosgenin or simvastatin by gavage every day for 8 weeks. The levels of triglyceride(TG), total cholesterol(TC), alanine transaminase(ALT), and aspartate transaminase(AST) in the serum were determined by the biochemical method. The levels of TG and TC in the liver were measured by the enzyme method. Oil-red O staining was employed to detect the lipid accumulation, and hematoxylin-eosin(HE) staining to detect the pathological changes in the liver tissue. The mRNA and protein levels of mTOR, SREBP-1c, HSP60, MCAD, and SCAD in the liver tissue of rats were determined by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR) and Western blot, respectively. Compared with the control group, the model group showed increased body weight, food uptake, liver index, TG, TC, ALT, and AST levels in the serum, TG and TC levels in the liver, lipid deposition in the liver, obvious hepatic steatosis, up-regulated mRNA and protein expression levels of mTOR and SREBP-1c, and down-regulated mRNA and protein expression levels of HSP60, MCAD, and SCAD. Compared with the model group, the rats in each treatment group showed obviously decreased body weight, food uptake, liver index, TG, TC, ALT, and AST levels in the serum, TG and TC levels in the liver, lessened lipid deposition in the liver, ameliorated hepatic steatosis, down-regulated mRNA and protein le-vels of mTOR and SREBP-1c, and up-regulated mRNA and protein levels of HSP60, MCAD, and SCAD. The high-dose diosgenin outperformed the low-dose diosgenin and simvastatin. Diosgenin may prevent and treat NAFLD by inhibiting the expression of mTOR and SREBP-1c and promoting the expression of HSP60, MCAD, and SCAD to reduce lipid synthesis, improving mitochondrial function, and promoting fatty acid ß oxidation in the liver.

Simvastatin Reduces Doxorubicin-Induced Cardiotoxicity: Effects beyond Its Antioxidant Activity.

This study aimed to evaluate if Simvastatin can reduce, and/or prevent, Doxorubicin (Doxo)-induced cardiotoxicity. H9c2 cells were treated with Simvastatin (10 µM) for 4 h and then Doxo (1 µM) was added, and the effects on oxidative stress, calcium homeostasis, and apoptosis were evaluated after 20 h. Furthermore, we evaluated the effects of Simvastatin and Doxo co-treatment on Connexin 43 (Cx43) expression and localization, since this transmembrane protein forming gap junctions is widely involved in cardioprotection. Cytofluorimetric analysis showed that Simvastatin co-treatment significantly reduced Doxo-induced cytosolic and mitochondrial ROS overproduction, apoptosis, and cytochrome c release. Spectrofluorimetric analysis performed by means of Fura2 showed that Simvastatin co-treatment reduced calcium levels stored in mitochondria and restored cytosolic calcium storage. Western blot, immunofluorescence, and cytofluorimetric analyses showed that Simvastatin co-treatment significantly reduced Doxo-induced mitochondrial Cx43 over-expression and significantly increased the membrane levels of Cx43 phosphorylated on Ser368. We hypothesized that the reduced expression of mitochondrial Cx43 could justify the reduced levels of calcium stored in mitochondria and the consequent induction of apoptosis observed in Simvastatin co-treated cells. Moreover, the increased membrane levels of Cx43 phosphorylated on Ser368, which is responsible for the closed conformational state of the gap junction, let us to hypothesize that Simvastatin leads to cell-to-cell communication interruption to block the propagation of Doxo-induced harmful stimuli. Based on these results, we can conclude that Simvastatin could be a good adjuvant in Doxo anticancer therapy. Indeed, we confirmed its antioxidant and antiapoptotic activity, and, above all, we highlighted that Simvastatin interferes with expression and cellular localization of Cx43 that is widely involved in cardioprotection.

Simvastatin Hydroxy Acid Fails to Attain Sufficient Central Nervous System Tumor Exposure to Achieve a Cytotoxic Effect: Results of a Preclinical Cerebral Microdialysis Study.

3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors were potent hits against a mouse ependymoma cell line, but their effectiveness against central nervous system tumors will depend on their ability to cross the blood-brain barrier and attain a sufficient exposure at the tumor. Among 3-hydroxy-3-methylglutaryl coenzyme A inhibitors that had activity in vitro, we prioritized simvastatin (SV) as the lead compound for preclinical pharmacokinetic studies based on its potential for central nervous system penetration as determined from in silico models. Furthermore, we performed systemic plasma disposition and cerebral microdialysis studies of SV (100 mg/kg, p.o.) in a murine model of ependymoma to characterize plasma and tumor extracellular fluid (tECF) pharmacokinetic properties. The murine dosage of SV (100 mg/kg, p.o.) was equivalent to the maximum tolerated dose in patients (7.5 mg/kg, p.o.) based on equivalent plasma exposure of simvastatin acid (SVA) between the two species. SV is rapidly metabolized in murine plasma with 15 times lower exposure compared with human plasma. SVA exposure in tECF was <33.8 ± 11.9 µg/l per hour, whereas the tumor to plasma partition coefficient of SVA was <0.084 ± 0.008. Compared with in vitro washout IC50 values, we did not achieve sufficient exposure of SVA in tECF to suggest tumor growth inhibition; therefore, SV was not carried forward in subsequent preclinical efficacy studies.

Evaluation of a method using high performance liquid chromatography with ultraviolet detection for the determination of statins in macromycetes of the genus Pleurotus cultivated by fermentation processes.

The applicability of high-performance liquid chromatography with ultraviolet light (HPLC-UV) for the determination of the presence of statins in macromycetes of the genus Pleurotus was analyzed. The fungi were obtained by liquid-state fermentation (LSF) using unconventional sources of carbon as substrates and solid-state fermentation (SSF) employing agro industrial wastes. Five statins were used as standards: lovastatin and simvastatin in the lactone form (LOVL and SIML), their corresponding hydro-acidic forms (LOVH and SIMH) and pravastatin (PRA). The following measures were evaluated: the linearity, accuracy and precision, detection limit (DL) and quantification limit (QL). The results demonstrated HPLC-UV to be an effective tool for detecting the presence of statins in extracts of LSF and SSF products. Likewise, it was hypothesized that the strains that were used for the study do not produce statins. This finding highlights the importance of continuing to evaluate other strains of the same genus by using techniques such as HPLC to first separate sufficient quantities of the compounds that were detected using the standard technique but that did not match the retention time (tR) of any of the standards used.

Effect of the traditional Chinese medicine tongxinluo on endothelial dysfunction rats studied by using urinary metabonomics based on liquid chromatography-mass spectrometry.

A urinary metabonomic method based on ultra-fast liquid chromatography coupled with ion trap-time of flight mass spectrometry (UFLC/MS-IT-TOF) was employed to study the preventive efficacy and the metabolic changes caused by simavastatin and the traditional Chinese medicine tongxinluo in endothelial dysfunction rats. Principal component analysis (PCA) was applied to study metabolic patterns of endothelial dysfunction rats and healthy control rats. 1-Methyladenosine, indoxyl sulfate, hippuric acid, riboflavin, coproporphyrin, and p-cresol glucuronide were identified as potential biomarkers, indicating that pathways of adenine, tryptophan, phenylalanine, riboflavin and porphyrin metabolism were disturbed in endothelial dysfunction rats. Applications of simvastatin and tongxinluo to endothelial dysfunction rats improved endothelial function according to the results of histopathology and measurements of endothelin-1 and nitric oxide. Metabonomic studies suggested that tongxinluo prevents endothelial dysfunction by regulating multiple metabolic pathways to their normal state, whereas simvastatin only altered selected metabolic pathways. This research demonstrated that metabonomics is a powerful and promising tool for disease investigation and the efficacy evaluation of complex traditional Chinese medicines.

[Influence of gingko extract on metabolism of simvastatin].

OBJECTIVE: To conclude the possible interaction between gingko extract and simvastatin, by studing the effect of gingko extract on vitro metabolism of simvastatin and involving critical metabolic enzyme, which can guide the clinicians to use them rationally (propose good guideline for rational using of the two medicine mentioned above). METHOD: Thirty-two female SD rats were randomly separated into 4 groups, including negative control group. High dose of gingko extract. Low dose of gingko extract and positive control group. All groups were administered for 10 days with stomach tube, and then the quantity of liver microsome protein, the activity of CYP3A were determined by spectrophotograph simvastatin was incubated with the liver microsome, and the effect of gingko extract on its metabolism was estimated by measuring the amount of simvastatin by HPLC. RESULT: Comparing with the negative contrast group: the quantity of liver microsome protein, the activity of CYP3A and the metabolism of simvastatin in positive control group were all increased markly (P < 0.05). In high dose group, the quantity of liver microsome protein and the activity of CYP3A were both increased significantly (P < 0.05), and the metabolism of simvastatin also accelerate obviously (P < 0.05). But in the low dose group significant distinction of every index was not found. CONCLUSION: High dose of gingko extract can induce the activity of CYP3A, and promote the metabolism of simvastatin, so the medical interaction should be focused when gingko extract is coadministered with simvastatin and other substracts of CYP3A.

Can in vitro metabolism-dependent covalent binding data in liver microsomes distinguish hepatotoxic from nonhepatotoxic drugs? An analysis of 18 drugs with consideration of intrinsic clearance and daily dose.

In vitro covalent binding assessments of drugs have been useful in providing retrospective insights into the association between drug metabolism and a resulting toxicological response. On the basis of these studies, it has been advocated that in vitro covalent binding to liver microsomal proteins in the presence and the absence of NADPH be used routinely to screen drug candidates. However, the utility of this approach in predicting toxicities of drug candidates accurately remains an unanswered question. Importantly, the years of research that have been invested in understanding metabolic bioactivation and covalent binding and its potential role in toxicity have focused only on those compounds that demonstrate toxicity. Investigations have not frequently queried whether in vitro covalent binding could be observed with drugs with good safety records. Eighteen drugs (nine hepatotoxins and nine nonhepatotoxins in humans) were assessed for in vitro covalent binding in NADPH-supplemented human liver microsomes. Of the two sets of nine drugs, seven in each set were shown to undergo some degree of covalent binding. Among hepatotoxic drugs, acetaminophen, carbamazepine, diclofenac, indomethacin, nefazodone, sudoxicam, and tienilic acid demonstrated covalent binding, while benoxaprofen and felbamate did not. Of the nonhepatotoxic drugs evaluated, buspirone, diphenhydramine, meloxicam, paroxetine, propranolol, raloxifene, and simvastatin demonstrated covalent binding, while ibuprofen and theophylline did not. A quantitative comparison of covalent binding in vitro intrinsic clearance did not separate the two groups of compounds, and in fact, paroxetine, a nonhepatotoxin, showed the greatest amount of covalent binding in microsomes. Including factors such as the fraction of total metabolism comprised by covalent binding and the total daily dose of each drug improved the discrimination between hepatotoxic and nontoxic drugs based on in vitro covalent binding data; however, the approach still would falsely identify some agents as potentially hepatotoxic.

[Using simvastatin in combination with chophytol at early stages of cholelithiasis in elderly patients].

In the treatment of 78 elderly patients with the gallstone disease were used chophytol and vasilip. To estimate the efficiency of treatment we used modern laboratory-instrumental, ultrasonic and biochemical methods of examinations. The examinations of indexs of fat exchange in bile and blood showed the communication of theirs disturbances. The application of chophytol and vasilip in the treatment of the first stage of gallstone disease is prove the positive changes of clinical symptoms of illness and lead to improvement of liver functional condition and biochemical composition of bile. These changes are cause to decrease of risk of formation the gallstones.

Glucuronidation of statins in animals and humans: a novel mechanism of statin lactonization.

The active forms of all marketed hydroxymethylglutaryl (HMG)-CoA reductase inhibitors share a common dihydroxy heptanoic or heptenoic acid side chain. In this study, we present evidence for the formation of acyl glucuronide conjugates of the hydroxy acid forms of simvastatin (SVA), atorvastatin (AVA), and cerivastatin (CVA) in rat, dog, and human liver preparations in vitro and for the excretion of the acyl glucuronide of SVA in dog bile and urine. Upon incubation of each statin (SVA, CVA or AVA) with liver microsomal preparations supplemented with UDP-glucuronic acid, two major products were detected. Based on analysis by high-pressure liquid chromatography, UV spectroscopy, and/or liquid chromatography (LC)-mass spectrometry analysis, these metabolites were identified as a glucuronide conjugate of the hydroxy acid form of the statin and the corresponding delta-lactone. By means of an LC-NMR technique, the glucuronide structure was established to be a 1-O-acyl-beta-D-glucuronide conjugate of the statin acid. The formation of statin glucuronide and statin lactone in human liver microsomes exhibited modest intersubject variability (3- to 6-fold; n = 10). Studies with expressed UDP glucuronosyltransferases (UGTs) revealed that both UGT1A1 and UGT1A3 were capable of forming the glucuronide conjugates and the corresponding lactones for all three statins. Kinetic studies of statin glucuronidation and lactonization in liver microsomes revealed marked species differences in intrinsic clearance (CL(int)) values for SVA (but not for AVA or CVA), with the highest CL(int) observed in dogs, followed by rats and humans. Of the statins studied, SVA underwent glucuronidation and lactonization in human liver microsomes, with the lowest CL(int) (0.4 microl/min/mg of protein for SVA versus approximately 3 microl/min/mg of protein for AVA and CVA). Consistent with the present in vitro findings, substantial levels of the glucuronide conjugate (approximately 20% of dose) and the lactone form of SVA [simvastatin (SV); approximately 10% of dose] were detected in bile following i.v. administration of [(14)C]SVA to dogs. The acyl glucuronide conjugate of SVA, upon isolation from an in vitro incubation, underwent spontaneous cyclization to SV. Since the rate of this lactonization was high under conditions of physiological pH, the present results suggest that the statin lactones detected previously in bile and/or plasma following administration of SVA to animals or of AVA or CVA to animals and humans, might originate, at least in part, from the corresponding acyl glucuronide conjugates. Thus, acyl glucuronide formation, which seems to be a common metabolic pathway for the hydroxy acid forms of statins, may play an important, albeit previously unrecognized, role in the conversion of active HMG-CoA reductase inhibitors to their latent delta-lactone forms.