Dual-targeting tigecycline nanoparticles for treating intracranial infections caused by multidrug-resistant Acinetobacter baumannii.

Multidrug-resistant (MDR) Acinetobacter baumannii (A. baumannii) is a formidable pathogen responsible for severe intracranial infections post-craniotomy, exhibiting a mortality rate as high as 71%. Tigecycline (TGC), a broad-spectrum antibiotic, emerged as a potential therapeutic agent for MDR A. baumannii infections. Nonetheless, its clinical application was hindered by a short in vivo half-life and limited permeability through the blood-brain barrier (BBB). In this study, we prepared a novel core-shell nanoparticle encapsulating water-soluble tigecycline using a blend of mPEG-PLGA and PLGA materials. This nanoparticle, modified with a dual-targeting peptide Aß11 and Tween 80 (Aß11/T80@CSs), was specifically designed to enhance the delivery of tigecycline to the brain for treating A. baumannii-induced intracranial infections. Our findings demonstrated that Aß11/T80@CSs nanocarriers successfully traversed the BBB and effectively delivered TGC into the cerebrospinal fluid (CSF), leading to a significant therapeutic response in a model of MDR A. baumannii intracranial infection. This study offers initial evidence and a platform for the application of brain-targeted nanocarrier delivery systems, showcasing their potential in administering water-soluble anti-infection drugs for intracranial infection treatments, and suggesting promising avenues for clinical translation.

Efficient signal sequence of mRNA vaccines enhances the antigen expression to expand the immune protection against viral infection.

The signal sequence played a crucial role in the efficacy of mRNA vaccines against virus pandemic by influencing antigen translation. However, limited research had been conducted to compare and analyze the specific mechanisms involved. In this study, a novel approach was introduced by substituting the signal sequence of the mRNA antigen to enhance its immune response. Computational simulations demonstrated that various signal peptides differed in their binding capacities with the signal recognition particle (SRP) 54 M subunit, which positively correlated with antigen translation efficiency. Our data revealed that the signal sequences of tPA and IL-6-modified receptor binding domain (RBD) mRNA vaccines sequentially led to higher antigen expression and elicited more robust humoral and cellular immune protection against the SARS-CoV-2 compared to the original signal sequence. By highlighting the importance of the signal sequence, this research provided a foundational and safe approach for ongoing modifications in signal sequence-antigen design, aiming to optimize the efficacy of mRNA vaccines.

Diagnostic Accuracy of Computerized Bowel Sound Analysis with Non-Invasive Devices for Irritable Bowel Syndrome: A Systematic Review and Meta-Analysis.

Objective To assess the diagnostic accuracy of bowel sound analysis for irritable bowel syndrome (IBS) with a systematic review and meta-analysis. Methods We searched MEDLINE, EMBASE, the Cochrane Library, Web of Science, and IEEE Xplore databases until September 2023. Cross-sectional and case-control studies on diagnostic accuracy of bowel sound analysis for IBS were identified. We estimated the pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, and diagnostic odds ratio with a 95% confidence interval (CI), and plotted a summary receiver operating characteristic curve and evaluated the area under the curve. Results Four studies were included. The pooled diagnostic sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, and diagnostic odds ratio were 0.94 (95% CI, 0.87－0.97), 0.89 (95% CI, 0.81－0.94), 8.43 (95% CI, 4.81－14.78), 0.07 (95% CI, 0.03－0.15), and 118.86 (95% CI, 44.18－319.75), respectively, with an area under the curve of 0.97 (95% CI, 0.95－0.98). Conclusions Computerized bowel sound analysis is a promising tool for IBS. However, limited high-quality data make the results' validity and applicability questionable. There is a need for more diagnostic test accuracy studies and better wearable devices for monitoring and analysis.

ProTICS reveals prognostic impact of tumor infiltrating immune cells in different molecular subtypes.

Different subtypes of the same cancer often show distinct genomic signatures and require targeted treatments. The differences at the cellular and molecular levels of tumor microenvironment in different cancer subtypes have significant effects on tumor pathogenesis and prognostic outcomes. Although there have been significant researches on the prognostic association of tumor infiltrating lymphocytes in selected histological subtypes, few investigations have systemically reported the prognostic impacts of immune cells in molecular subtypes, as quantified by machine learning approaches on multi-omics datasets. This paper describes a new computational framework, ProTICS, to quantify the differences in the proportion of immune cells in tumor microenvironment and estimate their prognostic effects in different subtypes. First, we stratified patients into molecular subtypes based on gene expression and methylation profiles by applying nonnegative tensor factorization technique. Then we quantified the proportion of cell types in each specimen using an mRNA-based deconvolution method. For tumors in each subtype, we estimated the prognostic effects of immune cell types by applying Cox proportional hazard regression. At the molecular level, we also predicted the prognosis of signature genes for each subtype. Finally, we benchmarked the performance of ProTICS on three TCGA datasets and another independent METABRIC dataset. ProTICS successfully stratified tumors into different molecular subtypes manifested by distinct overall survival. Furthermore, the different immune cell types showed distinct prognostic patterns with respect to molecular subtypes. This study provides new insights into the prognostic association between immune cells and molecular subtypes, showing the utility of immune cells as potential prognostic markers. Availability: R code is available at https://github.com/liu-shuhui/ProTICS.

Propagation of noninteger cylindrical vector vortex beams in a gradient-index fiber.

The characteristics of two noninteger cylindrical vector vortex beams (NCVVBs) propagating through a radial gradient-index (GRIN) fiber are analyzed on the basis of the generalized Huygens-Fresnel principle. The NCVVBs exhibit periodic and stable transmission characteristics in the radial GRIN fiber. Polarization changes, the presence of spin angular momentum (SAM), and changes in the orbital angular momentum (OAM) of the NCVVBs are observed at the focal plane of the radial GRIN fiber. Spin-orbit interactions of NCVVBs are verified in the radial GRIN fiber for the first time, to the best of our knowledge.

Facial Micro-Expression Recognition Using Double-Stream 3D Convolutional Neural Network with Domain Adaptation.

Humans show micro-expressions (MEs) under some circumstances. MEs are a display of emotions that a human wants to conceal. The recognition of MEs has been applied in various fields. However, automatic ME recognition remains a challenging problem due to two major obstacles. As MEs are typically of short duration and low intensity, it is hard to extract discriminative features from ME videos. Moreover, it is tedious to collect ME data. Existing ME datasets usually contain insufficient video samples. In this paper, we propose a deep learning model, double-stream 3D convolutional neural network (DS-3DCNN), for recognizing MEs captured in video. The recognition framework contains two streams of 3D-CNN. The first extracts spatiotemporal features from the raw ME videos. The second extracts variations of the facial motions within the spatiotemporal domain. To facilitate feature extraction, the subtle motion embedded in a ME is amplified. To address the insufficient ME data, a macro-expression dataset is employed to expand the training sample size. Supervised domain adaptation is adopted in model training in order to bridge the difference between ME and macro-expression datasets. The DS-3DCNN model is evaluated on two publicly available ME datasets. The results show that the model outperforms various state-of-the-art models; in particular, the model outperformed the best model presented in MEGC2019 by more than 6%.

Ethylene-induced potassium transporter AcKUP2 gene is involved in kiwifruit postharvest ripening.

BACKGROUND: Potassium (K) is important in the regulation of plant growth and development. It is the most abundant mineral element in kiwifruit, and its content increases during fruit ripening. However, how K+ transporter works in kiwifruit postharvest maturation is not yet clear. RESULTS: Here, 12 K+ transporter KT/HAK/KUP genes, AcKUP1 ~ AcKUP12, were isolated from kiwifruit, and their phylogeny, genomic structure, chromosomal location, protein properties, conserved motifs and cis-acting elements were analysed. Transcription analysis revealed that AcKUP2 expression increased rapidly and was maintained at a high level during postharvest maturation, consistent with the trend of K content; AcKUP2 expression was induced by ethylene, suggesting that AcKUP2 might play a role in ripening. Fluorescence microscopy showed that AcKUP2 is localised in the plasma membrane. Cis-elements, including DER or ethylene response element (ERE) responsive to ethylene, were found in the AcKUP2 promoter sequence, and ethylene significantly enhanced the AcKUP2 promoter activity. Furthermore, we verified that AcERF15, an ethylene response factor, directly binds to the AcKUP2 promoter to promote its expression. Thus, AcKUP2 may be an important potassium transporter gene which involved in ethylene-regulated kiwifruit postharvest ripening. CONCLUSIONS: Therefore, our study establishes the first genome-wide analysis of the kiwifruit KT/HAK/KUP gene family and provides valuable information for understanding the function of the KT/HAK/KUP genes in kiwifruit postharvest ripening.

mRNA Vaccines Against SARS-CoV-2 Variants Delivered by Lipid Nanoparticles Based on Novel Ionizable Lipids.

SARS-CoV-2 variants are now still challenging all the approved vaccines, including mRNA vaccines. There is an urgent need to develop new generation mRNA vaccines with more powerful efficacy and better safety against SARS-CoV-2 variants. In this study, a new set of ionizable lipids named 4N4T are constructed and applied to form novel lipid nanoparticles called 4N4T-LNPs. Leading 4N4T-LNPs exhibit much higher mRNA translation efficiency than the approved SM-102-LNPs. To test the effectiveness of the novel delivery system, the DS mRNA encoding the full-length S protein of the SARS-CoV-2 variant is synthesized and loaded in 4N4T-LNPs. The obtained 4N4T-DS mRNA vaccines successfully trigger robust and durable humoral immune responses against SARS-CoV-2 and its variants including Delta and Omicron. Importantly, the novel vaccines have higher RBD-specific IgG titers and neutralizing antibody titers than SM-102-based DS mRNA vaccine. Besides, for the first time, the types of mRNA vaccine-induced neutralizing antibodies are found to be influenced by the chemical structure of ionizable lipids. 4N4T-DS mRNA vaccines also induce strong Th1-skewed T cell responses and have good safety. This work provides a novel vehicle for mRNA delivery that is more effective than the approved LNPs and shows its application in vaccines against SARS-CoV-2 variants.

3D variable flip angle T1 mapping for differentiating benign and malignant liver lesions at 3T: comparison with diffusion weighted imaging.

BACKGROUND: Different methods have been used to improve the imaging diagnosis of focal liver lesions (FLL). Among them, magnetic resonance imaging (MRI) has received more attention since it provides significant amount of information without radiation exposure. However, atypical imaging characteristics of FLL on MRI may complicate the differential diagnosis between benign and malignant FLL. This study aimed to compare the diagnostic value of T1 mapping and diffusion-weighted imaging (DWI) for differentiating of benign and malignant FLLs. METHODS: This retrospective study enrolled 294 FLLs, including 150 benign and 144 malignant lesions. Whole liver T1 mapping sequences were obtained before and 2 min after the administration of Gd-DTPA to acquire native T1 and enhanced T1 and ΔT1%. Additionally, DWI sequence was conducted to generate apparent diffusion coefficient (ADC) maps. These quantitative parameters were compared using one-way analysis of variance, and the diagnostic accuracy of T1 mapping and ADC for FLLs was calculated by area under the curve (AUC). RESULTS: Significant differences were observed regarding the native T1, enhanced T1, ΔT1%, and ADC between benign and malignant FLLs. Furthermore, the sensitivity and specificity of the parameters are as follows: native T1 0.797/0.702 (cut off value 1635.5 ms); enhanced T1, 0.911/0.976 (cutoff value 339.2 ms); ΔT1%, 0.901/0.905 (cutoff value 70.8%); and ADC, 0.975/0.952 (cutoff value 1.21 × 10-3 mm2/s). The ideal cutoff values for native T1 and ADC in identifying cyst and haemangioma were 2422.9 ms (AUC 0.990, P < 0.01) and 2.077 × 10-3 mm2/s (AUC 0.949, P < 0.01), respectively, with a sensitivity and specificity of 0.963/1 and 0.852/0.892, respectively. ADC was significantly positively correlated with T1 and ΔT1%, and significantly negatively correlated with enhanced T1. CONCLUSION: The 3D Variable flip angle T1 mapping technique with Gd-DTPA has a high clinical potential for identifying benign and malignant FLLs. The enhanced T1 and ΔT1% values have similar diagnostic accuracy compared with DWI in evaluating FLLs. Native T1 shows better performance than DWI in distinguishing benign liver lesions, specifically, cysts, and haemangioma.

Prevention of ulcerative colitis by Huangqin decoction: reducing the intestinal epithelial cell apoptosis rate through the IFN-γ/JAK/ETS signalling pathway.

CONTEXT: Ulcerative colitis (UC) is a chronic idiopathic inflammatory bowel disease that is closely related to inflammation and apoptosis. The traditional Chinese medicine compound preparation Huangqin decoction (HQD) has been widely used in the clinical treatment of UC, but the specific mechanism of its function is still inconclusive. OBJECTIVE: To explore the pathogenesis of UC based on the IFN-γ/JAK/ETS signalling pathway, and to clarify the biological mechanism of HQD. MATERIALS AND METHODS: Forty 8-week-old male C57BL/6 mice were randomly divided into four groups: normal control, model, model + salazosulfapyridine group (500 mg/kg, p.o., pd) and model + HQD (9.1 g/kg, p.o., pd). Using Dextran sulphate sodium (DSS) salt (2.5%, p.o.)+high-fat diet + hot and humid environment to build a mouse model of UC. One month later, the changes of colon morphology, serum inflammatory factors, intestinal epithelial cell apoptosis and IFN-γ/JAK/ETS signalling pathway related protein changes in mice were observed. RESULTS: Compared with the model group, HQD significantly reduced the pathological score of the model mice's colon (2.60 ± 0.25 vs. 4.80 ± 0.37), and reduced the serum IFN-γ (200.30 ± 8.45 vs. 413.80 ± 6.97) and other inflammatory factors, and reduced intestinal epithelial cell apoptosis (24.85 ± 4.87 vs. 214.90 ± 39.21). In terms of mechanism, HQD down-regulated IFN-γ/JAK/ETS signalling pathway related proteins in colon tissue of UC model mice. CONCLUSIONS: These data indicate that HQD can improve UC by reducing intestinal inflammation and apoptosis, providing experimental evidence for the wide application of HQD in clinical practice of UC.

Saliency Detection with Moving Camera via Background Model Completion.

Detecting saliency in videos is a fundamental step in many computer vision systems. Saliency is the significant target(s) in the video. The object of interest is further analyzed for high-level applications. The segregation of saliency and the background can be made if they exhibit different visual cues. Therefore, saliency detection is often formulated as background subtraction. However, saliency detection is challenging. For instance, dynamic background can result in false positive errors. In another scenario, camouflage will result in false negative errors. With moving cameras, the captured scenes are even more complicated to handle. We propose a new framework, called saliency detection via background model completion (SD-BMC), that comprises a background modeler and a deep learning background/foreground segmentation network. The background modeler generates an initial clean background image from a short image sequence. Based on the idea of video completion, a good background frame can be synthesized with the co-existence of changing background and moving objects. We adopt the background/foreground segmenter, which was pre-trained with a specific video dataset. It can also detect saliency in unseen videos. The background modeler can adjust the background image dynamically when the background/foreground segmenter output deteriorates during processing a long video. To the best of our knowledge, our framework is the first one to adopt video completion for background modeling and saliency detection in videos captured by moving cameras. The F-measure results, obtained from the pan-tilt-zoom (PTZ) videos, show that our proposed framework outperforms some deep learning-based background subtraction models by 11% or more. With more challenging videos, our framework also outperforms many high-ranking background subtraction methods by more than 3%.

A network pharmacology-based approach to explore the effects of Chaihu Shugan powder on a non-alcoholic fatty liver rat model through nuclear receptors.

The pathogenesis of non-alcoholic fatty liver disease (NAFLD) is still not fully understood, and currently, no effective pharmacotherapy is available. Nuclear receptors (NRs) are important biological participants in NAFLD that exhibit great therapeutic potential. Chaihu Shugan powder (CSP) is a traditional Chinese medicine (TCM) formula that has a wide therapeutic spectrum including NAFLD, but the effective components and functional mechanisms of CSP are unclear. We adopted a network pharmacology approach using multiple databases for Gene Ontology (GO) enrichment analysis and the molecular complex detection (MCODE) method for a protein-protein interaction (PPI) analysis, and we used molecular docking method to screen the NR targets and determine the corresponding CSP components. The screening results were validated through a NAFLD rat model that was used to explain the possible relationship between CSP and NAFLD. Finally, we screened PPARγ, FXR, PPARα, RARα and PPARδ as target genes and quercetin, kaempferol, naringenin, isorhamnetin and nobiletin as target compounds. The five components were detected through high-performance liquid chromatography-mass spectrometry (HPLC-MS), the results of which aligned with the docking experiments of PPARγ, PPARα and PPARδ. After CSP intervention, the NAFLD rat model showed ameliorated effects in terms of bodyweight, hepatic histopathology, and serum and liver lipids, and the mRNA levels of PPARγ, FXR, PPARα and RARα were significantly changed. The results from this study indicate that CSP exhibits healing effects in an NAFLD model and that the network pharmacology approach to screening NR targets and determining the corresponding CSP components is a practical strategy for explaining the mechanism by which CSP ameliorates NAFLD.

Lipidomic Analysis of the Protective Effects of Shenling Baizhu San on Non-Alcoholic Fatty Liver Disease in Rats.

Shenling Baizhu San (SLBZS), a famous traditional Chinese medicine, has been demonstrated to exert protective effects against non-alcoholic fatty liver disease (NAFLD), but its exact mechanisms have not been well understood. The aim of this study was to investigate the mechanisms underlying the protective effects of SLBZS in a rat model of NAFLD using lipidomics and to evaluate the role of Sirtuin 1 (SIRT1) in the mechanism of SLBZS against NAFLD. The rat model of NAFLD was induced by high-fat feeding. An ultra-performance liquid chromatography-mass spectrometry (UHPLC-MS)-based untargeted lipidomics approach was applied to analyze hepatic lipid alterations, and the SIRT1-selective inhibitor EX 527 was used to inhibit SIRT expression in the liver. The results of body and biochemical parameters, as well as histological changes, indicated that SLBZS administration exerted protective effects against NAFLD. Lipidomic analysis showed that 30 lipid species were effectively regulated by SLBZS administration in rats fed a high-fat diet. Pathway analysis indicated that glycerophospholipid metabolism and glycerolipid metabolism were potential target pathways closely involved in the mechanism of SLBZS against NAFLD. Moreover, the beneficial effects of SLBZS on hepatic steatosis, some biochemical parameters and hepatic lipid species were partly diminished by SIRT1 inhibition. In conclusion, our results suggested that SLBZS administration could effectively alter some hepatic lipid species in rats fed a high-fat diet, which was mainly associated with the regulation of glycerophospholipid and glycerolipid metabolism. Furthermore, the beneficial effects of SLBZS on hepatic lipid metabolism may be at least partly attributed to SIRT1 activation in the liver.

A DNAzyme requiring two different metal ions at two distinct sites.

Most previously reported RNA-cleaving DNAzymes require only a single divalent metal ion for catalysis. We recently reported a general trivalent lanthanide-dependent DNAzyme named Ce13d. This work shows that Ce13d requires both Na(+) and a trivalent lanthanide (e.g. Ce(3+)), simultaneously. This discovery is facilitated by the sequence similarity between Ce13d and a recently reported Na(+)-specific DNAzyme, NaA43. The Ce13d cleavage rate linearly depends on the concentration of both metal ions. Sensitized Tb(3+) luminescence and DMS footprinting experiments indicate that the guanines in the enzyme loop are important for Na(+)-binding. The Na(+) dissociation constants of Ce13d measured from the cleavage activity assay, Tb(3+) luminescence and DMS footprinting are 24.6, 16.3 and 47 mM, respectively. Mutation studies indicate that the role of Ce(3+) might be replaced by G23 in NaA43. Ce(3+) functions by stabilizing the transition state phosphorane, thus promoting cleavage. G23 competes favorably with low concentration Ce(3+) (below 1 µM). The G23-to-hypoxanthine mutation suggests the N1 position of the guanine as a hydrogen bond donor. Together, Ce13d has two distinct metal binding sites, each fulfilling a different role. DNAzymes can be quite sophisticated in utilizing metal ions for catalysis and molecular recognition, similar to protein metalloenzymes.

Probiotic mixture of Lactobacillus and Bifidobacterium alleviates systemic adiposity and inflammation in non-alcoholic fatty liver disease rats through Gpr109a and the commensal metabolite butyrate.

AIMS: The study explored the systemic adiposity and inflammation through Gpr109a and the commensal metabolite butyrate during the treatment of non-alcoholic fatty liver disease rats with the probiotic mixture of Lactobacillus and Bifidobacterium for 16 weeks. METHODS: Fifteen male SD rats were randomly divided into three groups of five rats each: normal control group (basal feed), high-fat diet (HFD) feeding group (83% basal feed + 10% lard oil + 5% sucrose + 1.5% cholesterol + 0.5% cholate), and probiotic mixture intervention group (HFD + 0.6 g kg-1 day-1 probiotic mixture). Body composition, serum lipids, serum inflammatory markers, Gpr109a, and the commensal metabolite butyrate were assessed. RESULTS: Compared with HFD group, probiotic mixture significantly reduced body weight and the levels of serum FFA, TG, ALT, IL-1ß, and IL-18 (P < 0.05). The levels of Gpr109a and the commensal metabolite butyrate also changed significantly (P < 0.05). CONCLUSIONS: Probiotic mixture might inhibit systemic adiposity and inflammation through Gpr109a and the commensal metabolite butyrate in response to the insult of HFD.

PGK1 Drives Hepatocellular Carcinoma Metastasis by Enhancing Metabolic Process.

During the proliferation and metastasis, the tumor cells prefer glycolysis (Warburg effect), but its exact mechanism remains largely unknown. In this study, we demonstrated that phosphoglycerate kinase 1 (PGK1) is an important enzyme in the pathway of metabolic glycolysis. We observed a significant overexpression of PGK1 in hepatocellular carcinoma tissues, and a correlation between PGK1 expression and poor survival of hepatocellular carcinoma patients. Also, the depletion of PGK1 dramatically reduced cancer cell proliferation and metastasis, indicating an oncogenic role of PGK1 in liver cancer progression. Further experiments showed that PGK1 played an important role in MYC-induced metabolic reprogramming, which led to an enhanced Warburg effect. Our results revealed a new effect of PGK1, which can provide a new treatment strategy for hepatocellular carcinoma, as PGK1 is used to indicate the prognosis of hepatocellular carcinoma (HCC).

[Intervention of berberine on lipid deposition in liver cells of non-alcoholic fatty liver disease rats induced by high fat diet].

OBJECTIVE: To explore the effect of berberine on lipid metabolism disorder and lipid deposition in liver cells of non-alcoholic fatty liver disease (NAFLD) rats induced by high fat diet. METHODS: After one week adaptable feeding, 45 SPF level male SD rats were randomly divided into 3 groups, the normal control group, the model group, and the berberine group, 15 in each group. Except those in the normal control group, all rats were fed with high fat diet to prepare NAFLD model. As for rats in the berberine group, Berberine Hydrochloride was administered by gastrogavage. HE staining and oil red O staining were performed to identify the model after 8 weeks. Hepatocytes were isolated, and their activities and purities were tested by Typan blue staining and flow cytometry (FCM). Serum levels of TC, TG, HDL-C, and LDL-C were detected using automatic biochemical analyzer. mRNA expression levels of LXRα and FAS in liver cells were analyzed by Real-time quantitative polymerase chain reaction (PCR). Protein levels of LXRα and FAS in liver cells were examined by Western blot. RESULTS: The NAFLD rat model was successfully established by high fat diet. The yields of purified liver cells in each rat were (6.0-7.5) x 10(8). The viability of isolated liver cells with purity over 90% (tested by FCM analysis) was higher than 95%. Compared with the normal control group,the expression of LXRα and FAS at mRNA and protein levels was higher in the model group (P < 0.01). Compared with the model group, the expression of LXRα and FAS at mRNA and protein levels was obviously down-regulated in the berberine group (P < 0.01). CONCLUSIONS: LXRα/FAS signaling pathway was one of important signaling pathways of NAFLD lipid metabolism disorders. Berberine could recover hepatocyte fatty deposits in NAFLD rats by adjusting the LXR/FAS signaling pathway of hepatocytes, which might be one of important mechanisms for fighting against NAFLD.

Cilostazol promotes production of melanin by activating the microphthalmia-associated transcription factor (MITF).

Cilostazol, a licensed clinical drug for the treatment of intermittent claudication, is a phosphodiesterase (PDE) inhibitor that selectively inhibits PDE3, a cAMP-degrading enzyme, thus elevating levels of intracellular cAMP. It has been reported that pigment production by melanocytes both tans the skin and protects against skin cancers. The effects of cilostazol in melanogenesis are as yet unknown. In this study, treatment with cilostazol was found to promote the production of melanin as well as increase both Tyrosinase enzymatic activity and expression of the Tyrosinase gene. Importantly, we also found that cilostazol led to increased expression of the microphthalmia-associated transcription factor (MITF), the "master regulator" of both melanocyte differentiation and pigment production. Interestingly, knockdown of MITF using siRNA abolished the effects of cilostazol in melanogenesis, thereby suggesting that MITF might play a critical role in melanogenesis. Increased expression of MITF was abolished by treatment with H-89, a specific protein kinase A (PKA) inhibitor, thereby suggesting that the PKA pathway plays a critical role in cilostazol-induced expression of MITF. Cilostazol in fact enhanced expression of p-CREB, which was inhibited by H-89. Moreover, this cilostazol-induced increase in expression of MITF was inhibited by downregulation of CREB using CREB siRNA. These data suggest that induction of MITF via the PKA/CREB pathway plays a critical role in cilostazol-induced production of melanin in B16-F10 melanoma cells.

[Effect of Shugan Jianpi Recipe on LXRα/FAS signaling pathway mediated hepatocyte fatty deposits in NAFLD rats].

OBJECTIVE: To explore the effect of Shugan Jianpi Recipe (SJR) on LXRα/FAS signaling pathway mediated hepatocyte fatty deposits in nonalcoholic fatty liver disease (NAFLD) rats. METHODS: Totally 75 SPF grade male SD rats were randomly divided into 5 groups, i.e., the normal control group, the model group, the Shugan Recipe (SR) treatment groups, the Jianpi Recipe (JR) treatment group, and the SJR group. Except rats in the normal control group, the NAFLD rat model was duplicated using high fat diet (HFD). SR (Chaihu Shugan Powder) was administered to rats in the SR group. JR (Shenlin Baizhu Powder) was administered to rats in the JR group. SJR (Chaihu Shugan Powder plus Shenlin Baizhu Powder) was administered to rats in the SJR group. Changes of liver fat were analyzed using automatic biochemical analyzer. Liver cells were separated by low-speed centrifugation. Their activities and purities were identify using Typan blue and flow cytometry (FCM). Expression levels of LXRα and FAS mRNA in hepatocytes detected by Real-time quantitative PCR. Expression levels of LXRα and FAS protein were detected by Western blot. RESULTS: (1) Pathological results showed in the model group, hepatocytes were swollen with nucleus locating at the cell edge after oil red O staining; unequal sized small vacuoles could be seen inside cytoplasm. Some small vacuoles merged big vacuoles. All these indi- cated a NAFLD rat model was successfully established by high fat diet. Pathological structural changes could be impaired to some degree in all medicated groups, especially in the SR group. (2) Compared with the normal control group, expression levels of LXRα and FAS genes and proteins obviously increased in the model group (P < 0.01). Compared with the model group, their expression levels were obviously down-regulated in the JR group and the SR group (P < 0.01, P < 0.05). CONCLUSIONS: LXRα/FAS signaling pathway was an important signaling pathway for mediating lipid metabolism disorders of NAFLD rats. SJR could make hepatocyte fatty deposits tend to repair by adjusting the LXRα/FAS signaling pathway in NAFLD rats, which might be one of important mechanisms for SJR to prevent and cure NAFLD.

[Effects of soothing liver and invigorating spleen recipes on SREBP-1c, SCD-1 mRNA and proteins expression in hepatocytes of NAFLD rats].

OBJECTIVE: To observe the effects of soothing liver and invigorating spleen recipes on the expression levels of SREBP-1c, SCD-1 mRNA and proteins in hepatocytes of NAFLD rats,and to explore its possible mechanisms of prevention and treatment of NAFLD. METHODS: 75 SD male rats were randomly divided into 5 groups: normal group, model group, oothing liver group (administrated with 9.6 g/kg), invigorating spleen group (administrated with 30 g/kg)and integrated group (administrated with 39.6 g/kg). The rats of NASH model were induced by feeding a high-fat diet. After treatment for 8 weeks,9 rats were randomly taken to detect liver function, TC, TG and pathological changes in liver tissue. The other 6 rats of each group were taken respectively and collagenase (Type IV) was perfused to digest liver tissue with the circulation in vitro to separate hepatocytes. Real-time Q-PCR and Western Blot were used to detect the expression levels of SREBP-1c, SCD-1 mRNA and proteins. RESULTS: Compared with the model group,the different decrease levels of SREBP-1c, SCD-1 genes and proteins were found in all drug therapy groups (P < 0.05 or P < 0.01), as well different degrees that liver lipid and pathological changes became better, especially that of in soothing liver group. Comparison between the all drug therapy groups,the hepatocytes expression levels of SREBP-1c and SCD-1 mRNA in soothing liver group were lower than that of in invigorating spleen group (P < 0.05), but expression levels of the proteins had no statistical significances. CONCLUSION: Soothing liver and invigorating spleen recipes prevent and treat NAFLD,its mechanism may be related to inhibiting the activation of SREBP-1c/SCD-1 signal pathway in hepatocytes to down-regulate TC and TG synthesis and reduce hepatic lipid deposition. SREBP-1c, SCD-1 mRNA and proteins may be the effective targets.