Prevalence of porcine endogenous retroviruses in domestic, minature, and genetically modified pigs in Japan.

The present study examined the prevalence of porcine endogenous retroviruses (PERV) in pigs available in Japan using polymerase chain reaction (PCR) with primers specific for PERV-A, PERV-B, and PERV-C and for the full-length 5' to 3' long terminal repeat and using PCR-Southern blotting with env A-, env B-, env C-, and pol/pro-specific probes. All 376 pigs tested--Berkshire (B), Landrace (L), Duroc (D), Large White (W), miniature, and genetically modified triple-cross breed (LWD)--harbored both PERV-A and PERV-B genes. However, the prevalence of PERV-C differed among pigs: LWD, miniature, B, D, W, and L pigs were 100% (36/36), 83% (5/6), 68% (129/191), 52% (26/50), 21% (9/43), and 16% (8/50), respectively. These results show that W and L pigs may be preferable as xenotransplantation donors, because they may not produce human-tropic replication-competent hybrids of PERV-A and PERV-C.

Conceptual equivalence and health-related quality of life: an exploratory study in Japanese and Dutch cancer patients.

Research into the equivalence of Western and Japanese conceptualizations of health-related quality of life (HR-QOL) is scarce. We used the Western (European Organization for Research and Treatment of Cancer, EORTC-QLQ-C30) and the Japanese (HRQoL-20) questionnaire in order to analyze the conceptual similarity of HR-QOL factors, and the associations between specific symptom items with overall HR-QOL in Japanese (n=265) and Dutch (n=174) patients with various types of cancer. Both populations completed both instruments. In both patient groups, the overall health scale of the EORTC-QLQ-C30 correlated highly (r=0.59; p<0.001) with the HRQOL-20 composite average score, indicating substantial conceptual comparability. Relationships between all EORTC-QLQ-C30 symptom items with HR-QOL were examined by ranking their correlations with the two overall measures of HR-QOL. Comparable patterns in the Japanese and Dutch samples were observed. The results suggest a considerable conceptual equivalence of HR-QOL in Japanese and Dutch cancer patients, and indicate a satisfactory structural and cross-cultural equivalence for the EORTC-QLQ-C30 with regard to items measuring functioning and specific symptoms. Longitudinal studies are needed to examine the impact of specific symptoms on general quality of life.

Remodeling of the major pig xenoantigen by N-acetylglucosaminyltransferase III in transgenic pig.

We have been successful in generating several lines of transgenic mice and pigs that contain the human beta-d-mannoside beta-1,4-N-acetylglucosaminyltransferase III (GnT-III) gene. The overexpression of the GnT-III gene in mice and pigs reduced their antigenicity to human natural antibodies, especially the Galalpha1-3Galbeta1-4GlcNAc-R, as evidenced by immunohistochemical analysis. Endothelial cell studies from the GnT-III transgenic pigs also revealed a significant down-regulation in antigenicity, including Hanganutziu-Deicher antigen, and dramatic reductions in both the complement- and natural killer cell-mediated pig cell lyses. Changes in the enzymatic activities of other glycosyltransferases, such as alpha1,3-galactosyltransferase, GnT-IV, and GnT-V, did not support cross-talk between GnT-III and these enzymes in the transgenic animals. In addition, we demonstrated the effect of GnT-III in down-regulating the xenoantigen of pig heart grafts, using a pig to cynomolgus monkey transplantation model, suggesting that this approach may be useful in clinical xenotransplantation in the future.

Porcine MCP gene promoter directs high level expression of human DAF (CD55) in transgenic mice.

Porcine membrane cofactor protein (pMCP), a complement regulatory protein, is widely expressed in various tissues. Particularly, it is highly expressed on vascular endothelium. The objective of this study was to investigate whether the pMCP gene promoter can induce efficient expression of a human complement regulatory protein, decay-accelerating factor (DAF; CD55) in transgenic mice. Two fragments of the 5'-flanking region of pMCP gene (0.9 kb and 5.4 kb) connected with human DAF minigene (0.9/hDAF and 5.4/hDAF) were used to produce transgenic mice. The expression of hDAF in heart, liver, kidney, lung, pancreas, brain and testis of the transgenic mice was examined by immunohistochemical analysis. The vascular endothelia and the nerves in all organs examined were intensely stained. The staining pattern in these tissues was similar in all transgenic mice examined regardless of the length of the promoters. The surface expression levels of hDAF on peripheral red blood cells and splenocytes from a mouse carrying 5.4/hDAF hemizygously was twice the level of expression on corresponding human cells. The red blood cells and splenocytes from the transgenic mice exhibited resistance to lysis by human serum in a manner dependent upon expressed hDAF level. The hearts from the transgenic mice functioned for a significantly longer time than those from normal mice under perfusion with human serum in the Langendorff perfusion system. These results demonstrated that the pMCP gene promoter is a good candidate of the regulatory element in the transgene to produce transgenic animals for xenotransplantation.

Health-related quality of life varies with personality types: a comparison among cancer patients, non-cancer patients and healthy individuals in a Japanese population.

In an attempt to examine differential effects of personality on health-related quality of life (HRQoL) without regard to disease type, we used the HRQoL-20, a general questionnaire (Japanese original scale) we developed (comprising 20 questions related to physiological, psychological or social HRQoL) and the Eysenck Personality Questionnaire (EPQ), which measures personality traits of extraversion (E), neuroticism (N) and psychoticism (P). The subjects (399 males and 429 females), stomach cancer patients, non-cancer patients (who had received acupuncture or moxibustion treatment) and healthy controls, were classified into three personality types. The results indicated that the HRQoL score of the tolerable/tolerant type (high E, low N and high P scorers) was greater than the intolerable/intolerant type (low E, high N and low P scorers) and also the unclassified type (neither of above scorers). The HRQoL correlated positively with the E and P scales and negatively with the N scale, in the case of all subjects, with the exception of N in male cancer patients and E in male non-cancer patients. The results supported the hypothesis that the HRQoL varies with personality variables, in that each patient, in different treatment settings, strives for the situation that is congruent with his/her personality to attain a better HRQoL.

Intranasal administration of denatured type II collagen and its fragments can delay the onset of collagen-induced arthritis.

Collagen-induced arthritis (CIA) is an autoimmune animal model for some types of human rheumatoid arthritis (RA). We have evaluated the effectiveness of intranasal administration of antigen in inhibiting CIA in DBA/1 mice. The intranasal administration of heat-denatured or trypsin-digested bovine type II collagen (CII) before immunization with CII strongly delayed the onset of CIA, whereas administration of native CII did not do so. The mice administered denatured or digested CII possessed much lower titers of anti-CII IgG2a than the control mice, whereas titers of anti-CII IgG1 and IgG2b were unchanged or slightly decreased. Responding to CII and peptides containing immunodominant T cell determinants, lymph node cells from mice administered denatured CII produced less IFN-gamma. These results suggest that intranasal administration of antigen downregulated preferentially Th1-type responses, whereas an enhanced Th2-type response was not observed. We demonstrate that the methods shown here are a possible treatment for rheumatoid arthritis.

Molecular cloning of a pig homologue of membrane cofactor protein (CD46).

Organs of transgenic pigs that express human complement regulatory proteins are under assessment as an alternative to transplantation. A major barrier to the transplantation of pig organs is the hyperacute rejection caused by pre-existing antibodies and complement. Pig cells are very susceptible to human complement, presumably because pig cell-surface complement regulatory proteins are inefficient against it. Expression of human complement regulatory proteins, such as decay-accelerating factor and membrane cofactor proteins (MCP or CD46), by means of transgenes would confer resistance to human complement upon pig cells, thereby preventing hyperacute rejection. To express sufficient levels of human complement regulatory proteins at appropriate sites, regulatory elements of genes of pig membrane-bound complement regulatory proteins would be useful. To obtain their cDNAs, we transfected human cells with a pig cDNA library, selected cells by incubation with pig complement and rescued the plasmids. We cloned a cDNA for the pig homologue of MCP, pMCP. The cDNA encoded a predicted protein of 363 amino acids with 42% amino acid identity with human MCP. The pMCP consisted of four short consensus repeats, a Ser/Thr/Pro-rich domain, and transmembrane and cytoplasmic domains. Recombinant soluble pMCP that lacked transmembrane and cytoplasmic domains had factor I cofactor activity in C3b cleavage, indicating that it is functionally, as well as structurally homologous to MCP. FACS analysis with anti-pMCP mAb demonstrated that pMCP is expressed on all blood leukocytes, erythrocytes, and on endothelial and epithelial cell lines.

In vitro and in vivo anti-platelet effects of enzymatic hydrolysates of collagen and collagen-related peptides.

Collagen-related peptides, Gly-Pro-Arg and its analogues, were examined for their inhibitory effects on platelet aggregation induced by the addition of ADP. Human platelet aggregation was suppressed by more than 50% with each of Gly-Pro-Arg and such Gly-Pro-Arg-containing peptides as Gly-Pro-Arg-Gly, Gly-Pro-Arg-Gly-Pro, Gly-Pro-Arg-Pro-Pro, and Gly-Pro-Arg-Pro-Pro-Pro at a concentration of 0.3 mM. The inhibitory effects of these peptides were about 10 times higher in human PRP than in rat PRP. Other Gly-Pro-Arg analogues such as Sar-Pro-Arg, Gly-Pro-Lys, Gly-Ala-Arg, and Ala-Gly-Pro-Arg had no inhibitory effect at a concentration from 0.1 to 0.8 mM even in human PRP. Intravenous and oral administrations of Gly-Pro-Arg and enzymatic hydrolysates of collagen suppressed the decrease in platelet count for endotoxin-induced DIC in rats. Collagen itself has been regarded as a potent inducer of platelet aggregation, but these findings suggest that collagen-related peptides and enzymatic hydrolysates of collagen prevent platelet aggregation.

Histochemical and morphological studies on Trichinella spiralis larvae treated with high hydrostatic pressure.

Histochemical and morphological observations were made on Trichinella spiralis larvae treated with hydrostatic pressures of 100, 150, 200 and 300 MPa using hematoxylin-eosin (HE), periodic acid-Schiff (PAS) and Azan staining. Few histochemical changes were observed in HE and PAS stained larvae after pressurization at 200 MPa and under. However, red staining by Azan changed to blue in the anterior stichosome of larvae and skeletal muscle of mice, when the hydrostatic pressure was raised from 150 to 300 MPa. At 150 and 200 MPa, boundaries among stichocytes were indistinct or irregular, and unstained areas were observed in stichocytes of larvae using Azan staining. At 300 MPa, all tissues of larvae and mouse muscle stained blue with Azan. At the same pressure, decrease in PAS positive staining of stichocytes and dilation of muscular cells were observed in larvae. It is assumed that these histochemical and morphological changes in pressurized larvae might be related to the loss of infectivity of larvae.

Inactivation of herpes viruses by high hydrostatic pressure.

The effects of high hydrostatic pressure on herpes simplex virus type 1 (HSV-1) and human cytomegalovirus (HCMV) were examined. Pressure at more than 300 MPa for 10 min at 25 degrees C inactivated these virions and drastically inhibited their infection to cultured cells, and at greater than 400 MPa, reduced infective titers of HSV-1 and HCMV by more than 7 and 4 logs, respectively. Electron microscopic examination illustrated coincidentally that high pressure at 300 MPa damaged the virus envelope and prevented the virus particles from binding to the cells. The findings suggest that treatment at high hydrostatic pressure is promising as a means of inactivating HSV-1, HCMV and other enveloped viruses.

Reaction times and intelligence: a comparison of Japanese and British children.

Japanese and British 9-year-old children were compared on the standard progressive matrices and twelve reaction time parameters providing measures of simple and complex decision times, movement times and variabilities. The mean of the Japanese children on the progressive matrices exceeded that of the British children by 0.65 SD units and on the decision times component of reaction times by 0.50 SD units, suggesting that the high Japanese mean on psychometric intelligence is largely explicable in terms of the more efficient processing of information at the neurological level. Japanese children also showed faster movement times but, contrary to expectation, had greater variabilities than British children.

Effects of high hydrostatic pressure on characteristics of pork slurries and inactivation of microorganisms associated with meat and meat products.

Pork slurries inoculated with various test microorganisms were prepared and subjected to high hydrostatic pressure at 1000 to 6000 atm for 10 min at 25 degrees C to examine for the pressure effects on characteristics of the slurries and the inactivation of the microorganisms associated with meat and meat products. Pressure treatment at higher than 3000 atm caused coagulation and discoloration of the pork slurries. Harder and more white coagulants were obtained by increasing the pressure. Pressure treatment at 3000 to 6000 atm killed all the microorganisms tested by more than 6-log colony-forming units (cfu)/g except Bacillus cereus spores. Gram-negative microorganisms were more labile to pressure than Gram-positive ones. Campylobacter jejuni, Pseudomonas aeruginosa, Salmonella typhimurium and Yersinia enterocolitica were inactivated at pressures higher than 3000 atm; Escherichia coli, Saccharomyces cerevisiae and Candida utilis at pressures higher than 4000 atm; Micrococcus luteus, Staphylococcus aureus and Streptococcus faecalis at 6000 atm. Only less than one-log cfu/g of B. cereus spores were inactivated at 6000 atm. Ultraviolet absorption spectra and acridine orange staining suggested that E. coli became permeable and leaked cytoplasmic RNA at lower pressure than S. aureus. From the present findings, the authors propose high hydrostatic pressure treatment as a promising means of preparing wholesome meat and meat products.