Effect of chondroitin sulfate C on sperm capacitation and fertilization parameters in vitro in pigs.

The objective of this study was to determine the effects of chondroitin sulfate C (CS-C) on sperm capacitation and fertilization parameters in vitro in pigs. Frozen-thawed ejaculated pig sperm (semen S-484) were incubated with fertilization medium containing CS-C (0-2mg/ml) for 1h and the capacitation rate with chlorotetracycline (CTC) assay was examined, which showed that CS-C increased the rate of incapacitation F pattern spermatozoa converted to capacitation B pattern sperm cell in concentration-dependent manner and mostly increased capacitation B pattern sperm cell and decreased acrosome reaction AR pattern sperm cell in 1mg/ml concentration. When sperm was incubated for 1, 2 and 4h in fertilization medium containing 1mg/ml CS-C, it showed that the capacitated B pattern sperm cell was significantly (p<0.01) increased and the AR pattern sperm cell was significantly decreased at each time point in the presence than in the absence of CS-C. For identifying the validity of CS-C in sperm capacitation, sperm-oocyte was inseminated in fertilization medium containing CS-C (0-2mg/ml) and the rate of fertilized oocytes was examined, which showed that the penetration rates significantly (p<0.05) increased from 0.5 to 1.0mg/ml concentrations (87.4-96.3%) compared with control (74.9%). For identifying the universality of CS-C in sperm capacitation, four different semens (boar S-484, S-454, D-815 and D-748) were incubated in fertilization medium containing CS-C (1mg/ml) for 2h, respectively, which showed that CS-C increased the rate of capacitation B pattern sperm cell and decreased acrosome reaction AR pattern sperm cell in each semen. And it showed that CS-C yielded a higher promote effect (93.9%, 83.9%, 60.7% and 44.9%, respectively) on sperm penetration compared to unaddition control (63.4%, 22.0%, 3.3% and 3.3%, respectively). Sperm-oocyte binding analysis showed that CS-C increased the number of sperm bound to oocyte compared unaddition control in each semen. These results suggested that CS-C is the efficient factor on sperm capacitation in pigs, CS-C may promote sperm from the incapacitated to capacitated state and sequentially prevent sperm from spontaneous acrosome reaction, and thus facilitate the sperm-zona binding and sperm penetration to oocyte.

Keratitis, ichthyosis and deafness syndrome with development of multiple hair follicle tumours.

We report multiple occurrences of various kinds of tumours that originate from hair follicles in a patient diagnosed with KID (keratitis, ichthyosis, deafness) syndrome. These tumours are diagnosed as: (i) trichilemmal cysts in early lesions; (ii) proliferating trichilemmal tumours in moderate duration lesions; and (iii) malignant proliferating trichilemmal tumours in advanced lesions that are thought to progress from benign trichilemmal lesions. This three-step process raises the hypothesis of a multihit model that could account for the frequent development of tumours in KID patients.

[Bioinformatics analysis of autophagy 5 gene structure].

In order to study the relationship between autophagy and apoptosis, APG5 gene structure was revealed by bioinformatics analysis and meantime a new isoform resulted from alternative splicing of the hAPG5 gene was confirmed, which was hereby designated as human autophagy 5 beta (hAPG5 beta; LOCUS AF293841, GenBank). We cloned and sequenced the cDNAs from fetal brain and B cell cDNA libraries using the known hAPG5 cDNA open reading frame sequences as primers. The cDNA obtained from the human fetal brain cDNA library was identical to the known hAPG5 cDNA. However, the cDNA from adult brain cDNA library was 129 bp shorter in length, lacking the sequence corresponding to those from positions 434 to 563 of the hAPG5 cDNA. Through search in public database and sequence comparisons and assembly 4 related sequences, APG5 genomic sequence was obtained. We found that the hAPG5 gene had 8 exons, and those sequences missing in hAPG5 beta cDNA exactly corresponded to exon 3. By bioinformatics software, the structure of introns, exons, splicing sites, promoter and polyA were demonstrated. Moreover, we were able to express both hAGP5 and 5 beta cDNA clones in human hepatocytes and HeLa cells using pEGFP-C1 vector. In conclusion, our data indicate that a systematic bioinformatics method of finding protein diversity from alternative splicing is a good approach in post-genome biology.

Quantitative chromatics analysis for computer imaging of cytologic subtypes of lung cancer stained by Papanicolaou stain.

OBJECTIVE: To determine the role of quantitative chromatics analysis in the classification of subtypes of lung cancer stained by Papanicolaou stain. STUDY DESIGN: By means of computer image analysis, 60 keratinized squamous carcinoma cells (KSCC), 88 nonkeratinized squamous carcinoma cells (NKSCC) and 150 adenocarcinoma cells (ACC) from lung cancer in sputum smears stained by Papanicolaou stain were analyzed and distinguished based on quantitative colorimetry. The features measured were the content of three primary colors, red (R), green (G) and blue (B) and the coefficients of R, G and B (r, g and b, respectively). Hue, saturation, brightness and gray level were also measured. A stepwise discriminant analysis was carried out. RESULTS: The values of R, G and B and r, g and b, hue and saturation in NKSCC and ACC were significantly different from those of KSCC, and the changes in the three primary colors were more sensitive than those in the gray level. Computer assessment based on three primary color coefficients, hue and saturation yielded accuracy of distinguishing KSCC from NKSCC and KSCC from ACC of 95.2% and 95%, respectively. CONCLUSION: Quantitative analyses of R, G and B and r, g, b and hue and saturation are valuable in distinguishing KSCC from NKSCC and ACC.

[Effects of receptor-mediated endocytosis on macrophage membrane potential, cytosolic pH and lysosomal pH].

The effects of Con A, WGA, Zymosan A on membrane potential, cytosolic pH and lysosomal pH of cultured mouse macrophages were measured with fluorescence method. The results showed all the three kinds of ligands caused macrophage membrane potential hyperpolarization, cytosolic acidification and lysosomal alkalization, but reached their stabilized conditions at different times. The acidification of cytosl might inhibit further endocytosis. Alkalization of lysosomes was a basic factor for exocytosis of lysosomal contents. These receptor-mediated induced changes reflected cellular self-regulation and self-protection.

[Receptor-mediated endocytosis induced macrophage cytosolic acidification and exocytosis].

The effects of Con A, WGA, Zymosan A on macrophage cytosolic pH and outflow of lysosomal content through exocytosis were studied with SNAFL-calcein and FITC-Dextran on ACAS570. The results showed all three ligands could induce macrophage cytosolic acidification in about 10 min and kept at the same level hereafter; outflow of lysosomal fluorescent probe through exocytosis appeared in 15-20 min. In resting conditions, macrophage lysosomes mainly distributed in cell center; after stimulated for 15 min by three ligands, the number of lysosomes increased in membrane periphery, in 25-30 min lysosomes moved back toward cell center. We proposed that ligands induced lysosomal pH rises was a basic factor for outflow of lysosomal content through exocytosis, cytosolic acidification inhibited receptor-mediated endocytosis. Cytosolic acidification and outflow of lysosomal content through exocytosis were the results of cellular self-regulation and self-protection during receptor-mediated endocytosis.

[Ultrastructural observation of intratumoral neutrophils and macrophages induced by garlic oil].

After injection of garlic oil in tumor focus a large amount of neutrophils, macrophages and lymphocytes appeared. Some neutrophils and macrophages located adjacent to the tumor cells, some processes of neutrophils and macrophages penetrated into intracellular body of tumor cells. This result showed that garlic oil could induce neutrophils and macrophages against tumor.