Effects of various oil extraction methods on the gelatinization and retrogradation properties of starches isolated from tigernut (Cyperus esculentus) tuber meals.

Gelatinization and retrogradation characteristics of starches from tigernut (Cyperus esculentus) tuber before and after various oil extraction processes were studied in this investigation. The results indicated that starches isolated from tigernut tuber after the various oil extraction processes varied significantly in gelatinization and retrogradation properties. The starches isolated from the cakes of tigernut tuber after hot press extraction exhibited higher retrogradation tendency and relatively less shear-thinning than other starch samples. The results of FT-IR, XRD, and NMR analysis indicated that oil extraction had an unfavorable influence on starch retrogradation, which may be due to structural changes caused by oil extraction processes. In particular, oil extraction led to more efficient packing of double helices in the crystalline lamella of the starches during storage. Retrogradation of the starch gels also reduced the water holding capacities of the starches. The starch sample isolated from the cake after cold press extraction exhibited the highest water absorption capacity among the five samples for all storage times. This investigation provides valuable novel information for the industrial utilization of tigernut tuber starches isolated from meals and cakes after oil extraction.

N-acetylcysteine negatively regulates Notch3 and its malignant signaling.

Notch3 receptor is expressed in a variety of cancers and the excised active intracellular domain (N3ICD) initiates its signaling cascade. N-acetylcysteine (NAC) as an antioxidant has been implicated in cancer prevention and therapy. In this study, we demonstrated a negative regulation of Notch3 by NAC in cancer cells. HeLa cells treated with NAC exhibited a time- and concentration-dependent decrease in Notch3 levels and its downstream effectors Hes1 and HRT1 in a manner independent of f-secretase or glutathione. In contrast, NAC did not affect protein levels of Notch1, the full length Notch3 precursor, or ectopically expressed N3ICD. Although SOD, catalase and NAC suppressed reactive oxygen species in HeLa cells, the first two antioxidants did not impact on Notch3 levels. While the mRNA expression of Notch3 was not altered by NAC, functional inhibition of lysosome, but not proteasome, blocked the NAC-dependent reduction of Notch3 levels. Furthermore, results from Notch3 silencing and N3ICD overexpression demonstrated that NAC prevented malignant phenotypes through down-regulation of Notch3 protein in multiple cancer cells. In summary, NAC reduces Notch3 levels through lysosome-dependent protein degradation, thereby negatively regulates Notch3 malignant signaling in cancer cells. These results implicate a novel NAC treatment in sensitizing Notch3-expressing tumors.

Characteristics of testis-specific phosphoglycerate kinase 2 and its association with human sperm quality.

STUDY QUESTION: Is there an association between the expression of phosphoglycerate kinase (PGK) 2 in spermatozoa and sperm quality in both elderly men and young asthenozoospermia patients? SUMMARY ANSWER: Spermatozoa from elderly men and young asthenozoospermia patients show decreased expression of PGK2, which has a close positive relationship with sperm quality. WHAT IS KNOWN ALREADY: PGK1 and PGK2 are involved in spermatogenesis and thought to be related to sperm motility. However, limited information is known about their temporal-spatial expression in human spermatogenesis and their relationship with sperm quality. STUDY DESIGN, SIZE, DURATION: This was a case-control study including 30 healthy young males (aged 28-31 years), 30 elderly men (aged 68-70 years), and 30 asthenozoospermic patients (aged 25-40 years, progressive motility <32%) who donated semen samples. Furthermore, young testes samples were obtained from five fathers (27-33 years old) who had died in car accidents, while aged testes samples were obtained from five elderly fathers (78-82 years old) who were prostate cancer patients. PARTICIPANTS/MATERIALS, SETTING, METHODS: Semen samples from young adults, elderly men and asthenozoospermic patients were prepared, and their parameters were assessed by Computer-Aided Sperm Analysis (CASA). Sperm proteins were extracted for western blot analysis. Immunohistochemistry was used to characterize the cellular localization of PGK1 and PGK2 in testes samples. Sperm immunofluorescence quantification experiments identified the differential expression of PGK1 and PGK2 in sperm from young adults, elderly men and asthenozoospermic patients. Antibodies against PGK1 and PGK2 were used to test their influence on sperm motility and penetration into viscous media. A modified Kremer test using methyl cellulose was adopted to assess sperm function via penetration into viscous media. MAIN RESULTS AND THE ROLE OF CHANCE: Cellular localization analysis showed that PGK1 was mainly expressed in spermatogonia whereas PGK2 was mainly expressed in round spermatids. Expression levels of both PGKs were significantly decreased in the testis with ageing (P < 0.05). Western blot and immunofluorescence quantification showed markedly lower expression of PGK2 (P < 0.05) in sperm from elderly men or asthenozoospermic patients compared sperm from with healthy young men. Sperm functional analysis validated the close relationship between expression of PGK2 and sperm motility (staining percentage, r = 0.60, P < 0.05; intensity, r = 0.59, P < 0.05). Use of an anti-PGK2 antibody on sperm significantly decreased their ability to penetrate into a cervical mucus substitute (P < 0.05). LIMITATIONS, REASONS FOR CAUTION: Before any clinical applications using PGK2 to assess sperm quality can be developed, more cases should be used to evaluate this approach. WIDER IMPLICATIONS OF THE FINDINGS: The study provides new insights into the role of PGKs in male reproduction. The results also indicate that PGK2 is a promising molecular candidate for the assessment of sperm quality and the screening of male contraceptive targets. STUDY FUNDING/COMPETING INTERESTS: This work was supported by grants from the National Natural Science Foundation of China (no. 81300533, 81370013 and 81000277) and Shandong Provincial Natural Science Foundation, China (ZR2013HQ002). The authors declare no competing financial interests.

Comparison of gene expression of the oncogenic Wnt/ß-catenin signaling pathway components in the mouse and human epididymis.

ß-catenin is an integral part of the Wnt signaling pathway and has been linked to tumorigenesis and multiple developmental processes. The high ß-catenin expression with low tumor incidence in the human epididymis is thus intriguing. In the present study, the ß-catenin gene and protein was found to be highly expressed in the murine caput epididymidis, and the protein mainly localized along the lateral plasma membranes of adjacent epithelial cells throughout both human and mouse epididymides. Furthermore, the adult mouse epididymis was found to express almost all the Wnt/ß-catenin signaling pathway genes that were determined previously by our group in the human organ. Despite the differences in epididymal structure, the similar location of ß-catenin and the high concordance of this pathway's components' gene expression in both the adult human and mouse epididymides make the mouse a suitable animal model for studying the anti-tumor mechanism of the epididymis. In addition, both the mRNA and protein expression of ß-catenin shared a similar spatial expression as the mRNA of Ros1, a proto-oncogene and a key developmental regulator of the initial segment of the mouse epididymis. The observations on the parallel temporal expression of ß-catenin and Ros1 during postnatal development raise the possibility that the canonical Wnt signaling pathway has an additional role in the postnatal development of mouse epididymis.

Suppression of survivin expression by short hairpin RNA induces apoptosis in human laryngeal carcinoma cells.

PURPOSE: The aim of this study was to evaluate the suppression effect of survivin shRNA on the expression of the survivin gene in the human laryngeal cancer cell line Hep-2. PROCEDURES: 60 cases of laryngeal squamous-cell carcinoma (LSCC) and 10 cases of normal laryngeal mucosa were examined using immunohistochemistry to determine whether the expression of survivin correlated with tumorigenesis. Three plasmid vectors of short hairpin RNA (shRNA) specific for survivin were designed and generated. Western blot and real-time PCR analysis of survivin expression in Hep-2 cells was performed 48 h after transfection. The growth curve was used to determine the cell proliferation. Propidium iodide (PI) single staining was applied to detect the cell cycle. The apoptosis of the cells was analyzed by flow cytometry with the FITC-annexin-V/PI double staining and PI single staining. RESULTS: 68.33% (41 out of 60) of tumors were positive for survivin expression and significantly associated with lymph node metastasis and advanced stage. In contrast, no expression of survivin in normal mucosa was detected. Transfection of Hep-2 cells with survivin shRNA significantly inhibited survivin expression at both the mRNA and the protein level in Hep-2 cells. Downregulation of survivin resulted in increasing the apoptosis index, but the results showed no obvious influence on cell cycle. CONCLUSIONS: This study demonstrates that survivin shRNA effectively inhibits survivin gene expression in Hep-2 cells leading to growth suppression and apoptotic induction in Hep-2 cells.