From nutraceutical to clinical trial: frontiers in Ganoderma development

Ganoderma spp. are medical mushrooms with various pharmacological compounds which are regarded as a nutraceutical for improving health and treating diseases. This review summarizes current progress in the studies of Gamoderma ranging from bioactive metabolites, bioactivities, production techniques to clinical trials. Traditionally, polysaccharides and ganoderic acids have been reported as the major bioactive metabolites of Ganoderma possessing anti-tumor and immunomodulation functions. Moreover, recent studies indicate that Gandoerma also exerts other bioactivities such as skin lighting, gut microbiota regulation, and anti-virus effects. However, since these medical fungi are rare in natural environment, and that the cost of cultivation of fruiting bodies is high, industrial submerged fermentation of Ganoderma mycelia promotes the development of Ganoderma by dint of an increase of biomass and bioactive metabolites used for further application. In addition, various strategies for production of different metabolites are well developed, such as gene regulation, bi-stage pH, and oxygen control. To date, Ganoderma not only has become one of the most popular nutraceuticals worldwide but also has been applied to clinical trials for advanced diseases such as breast and non-small-cell lung cancer.

     

Colorectal cancer-associated fibroblasts promote metastasis by up-regulating LRG1 through stromal IL-6/STAT3 signaling

Cancer-associated fibroblasts (CAFs) have been shown to play a strong role in colorectal cancer metastasis, yet the underlying mechanism remains to be fully elucidated. Using CRC clinical samples together with ex vivo CAFs-CRC co-culture models, we found that CAFs induce expression of Leucine Rich Alpha-2-Glycoprotein 1(LRG1) in CRC, where it shows markedly higher expression in metastatic CRC tissues compared to primary tumors. We further show that CAFs-induced LRG1 promotes CRC migration and invasion that is concomitant with EMT (epithelial-mesenchymal transition) induction. In addition, this signaling axis has also been confirmed in the liver metastatic mouse model which displayed CAFs-induced LRG1 substantially accelerates metastasis. Mechanistically, we demonstrate that CAFs-secreted IL-6 (interleukin-6) is responsible for LRG1 up-regulation in CRC, which occurs through a direct transactivation by STAT3 following JAK2 activation. In clinical CRC tumor samples, LRG1 expression was positively correlated with CAFs-specific marker, α-SMA, and a higher LRG1 expression predicted poor clinical outcomes especially distant metastasis free survival, supporting the role of LRG1 in CRC progression. Collectively, this study provided a novel insight into CAFs-mediated metastasis in CRC and indicated that therapeutic targeting of CAFs-mediated IL-6-STAT3-LRG1 axis might be a potential strategy to mitigate metastasis in CRC.Subject terms: Colon cancer, Cancer microenvironment     

Cardiac mitochondrial ATP-sensitive potassium channel is activated by nitric oxide in vitro

Previous observations on the activation of the mitochondrial ATP-sensitive potassium channel (mitoK(ATP)) by nitric oxide (NO) in myocardial preconditioning were based on indirect evidence. In this study, we have investigated the direct effect of NO on the rat cardiac mitoK(ATP) after reconstitution of the inner mitochondrial membranes into lipid bilayers. We found that the mitoK(ATP) was activated by exogenous NO donor S-nitroso-N-acetyl penicillamine or PAPA NONOate. This activation was inhibited by mitoK(ATP) blockers 5-hydroxydecanoate or glibenclamide. Our observations confirm that NO can directly activate the cardiac mitoK(ATP), which may underlie its contribution to myocardial preconditioning.     

The use of synthetic genes for the expression of ciliate proteins in heterologous systems

The common fish parasite, Ichthyophthirius multifiliis, expresses abundant glycosylated phosphatidylinositol (GPI)-anchored membrane proteins known as immobilization antigens, or i-antigens. These proteins are targets of the host immune response, and have been identified as potential candidates for recombinant subunit vaccine development. Nevertheless, because Ichthyophthirius utilizes a non-standard genetic code, expression of the corresponding gene products, either as subunit antigens in conventional protein expression systems, or as vector-encoded antigens in the case of DNA vaccines, is far from straightforward. To overcome this problem, we utilized 'assembly polymerase chain reaction' to manufacture synthetic versions of two genes (designated IAG52A[G5/CC] and IAG52B[G5/CC]) encoding approximately 52/55 kDa i-antigens from parasite strain G5. This approach made it possible to eliminate unwanted stop codons and substitute the preferred codon usage of channel catfish for the native sequences of the genes. To determine whether the synthetic alleles could be expressed in cells that use the standard genetic code, we introduced IAG52A[G5/CC] into a variety of heterologous cell types and tested for expression either by immunofluorescence light microscopy or Western blotting. When cloned downstream of appropriate promoters, IAG52A[G5/CC] was expressed in Escherichia coli, mammalian COS-7 cells, and channel catfish where it elicited antigen-specific immune responses. Interestingly, the localization pattern of the corresponding gene product in COS-7 cells indicated that while the protein was correctly folded, it was not present on the cell membrane, suggesting that the signal peptides required for GPI-anchor addition differ in ciliate and mammalian systems. Construction of synthetic alleles should have practical utility in the development of vaccines against Ichthyophthirius, and at the same time, provide a general method for the expression of ciliate genes in heterologous systems.     

Metabolomic approach to optimizing and evaluating antibiotic treatment in the axenic culture of cyanobacterium Nostoc flagelliforme

The application of antibiotic treatment with assistance of metabolomic approach in axenic isolation of cyanobacterium Nostoc flagelliforme was investigated. Seven antibiotics were tested at 1–100 mg L^?1, and order of tolerance of N. flagelliforme cells was obtained as kanamycin > ampicillin, tetracycline > chloromycetin, gentamicin > spectinomycin > streptomycin. Four antibiotics were selected based on differences in antibiotic sensitivity of N. flagelliforme and associated bacteria, and their effects on N. flagelliforme cells including the changes of metabolic activity with antibiotics and the metabolic recovery after removal were assessed by a metabolomic approach based on gas chromatography–mass spectrometry combined with multivariate analysis. The results showed that antibiotic treatment had affected cell metabolism as antibiotics treated cells were metabolically distinct from control cells, but the metabolic activity would be recovered via eliminating antibiotics and the sequence of metabolic recovery time needed was spectinomycin, gentamicin > ampicillin > kanamycin. The procedures of antibiotic treatment have been accordingly optimized as a consecutive treatment starting with spectinomycin, then gentamicin, ampicillin and lastly kanamycin, and proved to be highly effective in eliminating the bacteria as examined by agar plating method and light microscope examination. Our work presented a strategy to obtain axenic culture of N. flagelliforme and provided a method for evaluating and optimizing cyanobacteria purification process through diagnosing target species cellular state.     

尿激酶胸腔注入治疗结核包囊性胸腔积液

目的寻求治疗结核包囊性胸腔积液的新方法。方法胸膜活检确诊为结核性胸腔积液并经Ｂ超或胸部Ｘ线证实有粘连包囊的病人１５例,经胸腔内注入尿激酶每次１０万单位,用６０ｍｌ生理盐水稀释,每８～１２ｈ重复,连续５次,每次记录抽取胸水总量,并复查Ｂ超观察粘连包囊情况。对照组为单纯用抗结核治疗及间断抽胸水１５例。结果实验组１３例有效,总有效率８６．７％。对照组２例有效,有效率１３．３％,疗效明显优于对照组（Ｐ＜０．０５）。结论尿激酶胸腔内注入是治疗结核性粘连包囊性胸腔积液的一种安全可靠的新方法。