Molecular targeting for the altered signaling pathways has been proven to be effective for the treatment of many types of human cancer, including colorectal cancer (CRC). The dual phosphatidylinositol-3-kinase (PI3K) and mammalian target of rapamycin (mTOR) inhibitor BEZ235 has shown to exhibit potent antitumor activity against solid tumors. Autophagy is a cellular lysosomal catabolic process to maintain metabolic homeostasis, which has been known to be induced in response to many therapeutic agents in cancer cells. This process is negatively regulated by mTOR and often acts as prosurvival or prodeath mechanism following cancer therapeutics. The current study was designed to investigate the antiproliferation activity of BEZ235 and to evaluate the role of autophagy induced by BEZ235 using HCT15 CRC cells bearing ras oncogene mutation. We found that BEZ235 decreases cell viability, which was mostly dependent on G1 arrest of cell cycle via suppression of cyclin A expression. BEZ235 affects PI3K/Akt/mTOR signaling pathway by increasing the phosphorylation of AKT at Ser473 and RAS/RAF/MEK/ERK pathway by decreasing the phosphorylation of ERK at Tyr204. BEZ235 also stimulated autophagy induction as evidenced by the increased expression of LC3-II and abundant acidic vesicular organelles (AVOs) in the cytoplasm. In addition, the combination of BEZ235 with autophagy inhibitor chloroquine, a known antagonist of autophagy, counteracted the antiproliferation effect of BEZ235. Thus, our study indicates that autophagy induced in response to BEZ235 treatment appears to act as cell death mechanism in HCT15 CRC cells.
The ultrastructures of germ cells and the functions of Leydig cells and Sertoli cells during spermatogenesis in male Kareius bicoloratus (Pleuronectidae) were investigated by electron microscope observation. Each of the well-developed Leydig cells during active maturation division and before spermiation contained an ovoid vesicular nucleus, a number of smooth endoplasmic reticula, well-developed tubular or vesicular mitochondrial cristae, and several lipid droplets in the cytoplasm. It is assumed that Leydig cells are typical steroidogenic cells showing cytological characteristics associated with male steroidogenesis. No cyclic structural changes in the Leydig cells were observed through the year. However, although no clear evidence of steroidogenesis or of any transfer of nutrients from the Sertoli cells to spermatogenic cells was observed, cyclic structural changes in the Sertoli cells were observed over the year. During the period of undischarged germ cell degeneration after spermiation, the Sertoli cells evidenced a lysosomal system associated with phagocytic function in the seminiferous lobules. In this study, the Sertoli cells function in phagocytosis and the resorption of products originating from degenerating spermatids and spermatozoa after spermiation. The spermatozoon lacks an acrosome, as have been shown in all teleost fish spermatozoa. The flagellum or sperm tail of this species evidences the typical 9+2 array of microtubules.
In fish, light (photoperiod, intensity and spectra) is main regulator in many physiological actions including growth. We investigate the effect of light spectra on the somatic growth and growth-related gene expression in the rearing tiger puffer. Fish was reared under different light spectra (blue, green and red) for 8 weeks. Fish body weight and total length were promoted when reared under green light condition than red light condition. Expression of somatostatins (ss1 and ss2) in brain were showed higher expression under red light condition than green light condition. The ss3 mRNA was observed only higher expression in blue light condition. Expression of growth hormone (gh) in pituitary was detected no different levels between experimental groups. However, the fish of green light condition group was showed more high weight gain and feed efficiency than other light condition groups. Our present results suggest that somatic growth of tiger puffer is induced under green light condition because of inhibiting ss mRNA expression in brain by effect of green wavelength.
This study was conducted in order to examine the egg development in red spotted grouper, Epinephelus akaara and the morphological development of its larvae and juveniles, and to obtain data for taxonomic research. This study was conducted in June 2013, and 50 male and female fish were used for the study. One hundred μg/kg of LHRHa was injected into the body of the fish for inducing spawning, and the fish were kept in a small-sized fish holder (2×2×2 m). Eggs were colorless transparent free pelagic eggs, 0.71–0.77 mm large (mean 0.74±0.02 mm, n=30), and had an oil globule. Hatching started within 27 h after fertilization. Pre-larvae that emerged just after hatching were 2.02–2.17 mm in total length (mean 2.10±0.11 mm), their mouth and anus were not opened yet, and the whole body was covered with a membrane fin. Post-larvae that emerged 15 days post hatching were 3.88–4.07 mm in total length (mean 3.98±0.13 mm), and had a ventral fin with two rays and a caudal fin with eight rays. Juveniles that were formed at 55 d post hatching, were 31.9–35.2 mm in total length (mean 33.6±2.33 mm), with red color deposited over the entire body, and black chromophores deposited in a spotted pattern. The number of fin rays, body color, and shape were the same as that in the adult fish.
Neurokinin B (NKB) and neurokinin B related peptide (NKBRP) belong to tachykinin peptide family. They act as a neurotransmitter and/or neuromodulator. Mutation of NKB and/or its cognate receptor, NK3R resulted in hypogonadotropic hypogonadism in mammals, implying a strong involvement of NKB/NK3R system in controlling mammalian reproduction. Teleosts possess NKBRP as well as NKB, but their roles in fish reproduction need to be clarified. In this study, NKB and NKBRP coding gene (tac3) was cloned from Nile tilapia and sequenced. Based on the sequence, Nile tilapia NKB and NKBRP peptide were synthesized and their biological potencies were tested in vitro pituitary culture. The synthetic NKBRP showed direct inhibitory effect on the expression of GTH subunits at the pituitary level. This inhibitory effect was confirmed in vivo by means of intraperitoneal (ip) injection of synthetic NKB and NKBRP to mature female tilapia (20 pmol/g body weight [BW]). Both NKB and NKBRP had no effect on the plasma level of sex steroids, E2 and 11-KT. However, NKBRP caused declines of expression level of GnRH I, Kiss2 and tac3 mRNAs in the brain while NKB seemed to have no distinct effect. These results indicate some inhibitory roles of NKBRP in reproduction of mature female Nile tilapia, although their exact functions are not clear at the moment.
Human embryonic stem cells (hESCs) have been routinely cultured on mouse embryonic fibroblast feeder layers with a medium containing animal materials. For clinical application of hESCs, animal-derived products from the animal feeder cells, animal substrates such as gelatin or Matrigel and animal serum are strictly to be eliminated in the culture system. In this study, we performed that SNUhES32 and H1 were cultured on human amniotic fluid cells (hAFCs) with KOSR XenoFree and a humanized substrate. All of hESCs were relatively well propagated on hAFCs feeders with xeno-free conditions and they expressed pluripotent stem cell markers, alkaline phosphatase, SSEA-4, TRA1-60, TRA1-81, Oct-4, and Nanog like hESCs cultured on STO or human foreskin fibroblast feeders. In addition, we observed the expression of nonhuman N-glycolylneuraminic acid (Neu5GC) molecules by flow cytometry, which was xenotransplantation components of contamination in hESCs cultured on animal feeder conditions, was not detected in this xeno-free condition. In conclusion, SNUhES32 and H1 could be maintained on hAFCs for humanized culture conditions, therefore, we suggested that new xenofree conditions for clinical grade hESCs culture will be useful data in future clinical studies.