Effects of Graphene Oxide Nanofilm and Chicken Embryo Muscle Extract on Muscle Progenitor Cell Differentiation and Contraction

Finding an effective muscle regeneration technique is a priority for regenerative medicine. It is known that the key factors determining tissue formation include cells, capable of proliferating and/or differentiating, a niche (surface) allowing their colonization and growth factors. The interaction between these factors, especially between the surface of the artificial niche and growth factors, is not entirely clear. Moreover, it seems that the use of a complex of complementary growth factors instead of a few strictly defined ones could increase the effectiveness of tissue maturation, including muscle tissue. In this study, we evaluated whether graphene oxide (GO) nanofilm, chicken embryo muscle extract (CEME), and GO combined with CEME would affect the differentiation and functional maturation of muscle precursor cells, as well as the ability to spontaneously contract a pseudo-tissue muscle. CEME was extracted on day 18 of embryogenesis. Muscle cells obtained from an 8-day-old chicken embryo limb bud were treated with GO and CEME. Cell morphology and differentiation were observed using different microscopy methods. Cytotoxicity and viability of cells were measured by lactate dehydrogenase and Vybrant Cell Proliferation assays. Gene expression of myogenic regulatory genes was measured by Real-Time PCR. Our results demonstrate that CEME, independent of the culture surface, was the main factor influencing the intense differentiation of muscle progenitor cells. The present results, for the first time, clearly demonstrated that the cultured tissue-like structure was capable of inducing contractions without externally applied impulses. It has been indicated that a small amount of CEME in media (about 1%) allows the culture of pseudo-tissue muscle capable of spontaneous contraction. The study showed that the graphene oxide may be used as a niche for differentiating muscle cells, but the decisive influence on the maturation of muscle tissue, especially muscle contractions, depends on the complexity of the applied growth factors.

Bovine fetal tissue extracts as an alternative to fetal serum for in vitro reproduction of viruses

The authors assessed the possibility of substitution of the serum component with tissue extracts (muscles, liver, kidneys) of bovine fetuses in the culture medium during the cultivation of transplanted LEK and Vero cell lines, as well as the reproduction of infectious rhinotracheitis IR, PI-3 viruses and reovirus on them. The greatest stimulating effect on LEK and Vero cells was obtained from bovine fetuses muscle extract. The effect of this extract on the proliferative activity of LEK and Vero cells is significant and amounts to 27 and 25%, respectively. The power of the effect of liver and kidney extracts is significantly lower and equal, respectively, 15 and 18% for LEK and 14 and 19% for Vero, although it is reliable. The reproductive activity of IR and PI-3 viruses when using tissue extracts was inferior to that when using blood serum. The stimulating effect of blood serum and muscle extract on the reproduction of reovirus was comparable. The effect of fetal muscle extract on the reproduction of IR, PI-3 viruses and reovirus is reliable and amounts to 29, 31 and 33%, respectively. In general, it is close to that of the blood serum of bovine fetuses - 30, 35 and 36%. The power of the influence of the liver and kidney extracts of the bovine fetuses is significantly lower and comparable to that of the blood serum of the cows themselves: 25, 23 and 20%, although it is reliable.

Dietary Supplementation of Selenoneine-Containing Tuna Dark Muscle Extract Effectively Reduces Pathology of Experimental Colorectal Cancers in Mice

Selenoneine is an ergothioneine analog with greater antioxidant activity and is the major form of organic selenium in the blood, muscles, and other tissues of tuna. The aim of this study was to determine whether a selenoneine-rich diet exerts antioxidant activities that can prevent carcinogenesis in two types of colorectal cancer model in mice. We administrated selenoneine-containing tuna dark muscle extract (STDME) to mice for one week and used azoxymethane (AOM) and dextran sodium sulfate (DSS) for inducing colorectal carcinogenesis. Next, we examined the incidence of macroscopic polyps and performed functional analysis of immune cells from the spleen. We also studied tumor formation rates and median survival following the subcutaneous implantation of a colorectal cancer cell line. In the AOM/DSS-induced colitis-associated cancer (CAC) model, the oral administration of STDME significantly decreased tumor incidence and inhibited the accumulation of myeloid-derived suppressor cells (MDSCs) while also inhibiting the downregulation of interferon-γ (IFN-γ) production during carcinogenesis. These results suggest that dietary STDME may be an effective agent for reducing colorectal tumor progression.

Tetrodotoxin Detection by a Surface Plasmon Resonance Sensor in Pufferfish Matrices and Urine

Tetrodotoxin (TTX) poisoning is most commonly associated with consumption of pufferfish. TTX is a low molecular weight (~319 Da) neurotoxin that selectively blocks voltage-sensitive Na+-gated ion channels. The standard method accepted worldwide for monitoring TTX toxicity in food matrices is the mouse bioassay. Ethical concerns from live animal testing, low sample throughput, and analytical inaccuracies have led to the need for an alternative method. We have previously established that surface plasmon resonance (SPR) sensors can quantify TTX in aqueous buffer samples by an antibody-based inhibition assay. In this paper, we report the extension of the assay for the detection of TTX in both clinical- and food-relevant matrices. The assay was optimized for application to three relevant complex matrices: pufferfish liver extract, pufferfish muscle extract, and human urine. Matrix effects are discussed and calibration curves are presented. Naturally contaminated pufferfish liver and muscle extracts were analyzed by the SPR method, and the data is compared to liquid-chromatography electrospray-ionization multiple reactions monitoring mass spectrometry (LC/ESI/MRM/MS) data. Ten samples, including three from a poisoning incident, two control monkfish samples, and five toxic pufferfish samples, were analyzed using this method, and the data is compared to LC/ESI/MRM/MS analysis of the samples.