• Journal of Embryo Transfer
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• v.12 no.3
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• pp.247-251
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• 1997

This research was conducted to observe developmental capacity of the early embryos aggrigated to phytohemagglutinin-M(PHA-M) in the culture of mouse embryos in vitro. The results showed that the development of blastocyst increased to 2-celT >< 2-cell : 68. 9%, 4-cell $\times$4-cell : 92.5% and 8-cell $\times$8-cell : 97.3% in the aggrigated embryos of ICR mouse, and 2-cell $\times$ 2-cell : 90.0%, 4-cell $\times$4-cell : 93.9% and 8-cell $\times$ 8-cell : 100% in the aggrigated embryos of two different strains (ICR $\times$ CBA/J mouse). (Key words : aggrigated embryos, in vitro 2-cell block, phytohemagglutinin-M, blastocyst)

• BMB Reports
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• v.35 no.5
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• pp.472-475
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• 2002

Phaseolus vulgaris phytohemagglutinin L is a homotetrameric-leucoagglutinating seed lectin. Its three-dimensional structure shows similarity with other members of the legume lectin family. The tetrameric form of this lectin is pH dependent. Gel filtration results showed that the protein exists in its dimeric state at pH 2.5 and as a tetramer at pH 7.2. Contrary to earlier reports on legume lectins that possess canonical dimers, thermal denaturation studies show that the refolding of phytohemagglutinin L at neutral pH is irreversible. Differential scanning calorimetry (DSC) was used to study the denaturation of this lectin as a function of pH that ranged from 2.0 to 3.0. The lectin was found to be extremely thermostable with a transition temperature around $82^{\circ}C$ and above $100^{\circ}C$ at pH 2.5 and 7.2, respectively. The ratio of calorimetric to vant Hoff enthalpy could not be calculated because of its irreversible-folding behavior. However, from the DSC data, it was discovered that the protein remains in its compact-folded state, even at pH 2.3, with the onset of denaturation occurring at $60^{\circ}C$.

• Archives of Pharmacal Research
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• v.3 no.1
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• pp.31-36
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• 1980

Seeds or beans of 55 plants belonging to 31 families were screened by using several different types of red blood cells to find new lectins. In this paper, white kidney been (phaseolus vulgaris C.) was chosen to study biochemical properties of hemagglutinating proteins(lectins). An anion exchanger, DEAE Sephadex A-50, and polyacrylamide disc gel electrophoresis were main techniques used. From three main fractions eluted by stepwise NaCl gradient in 25mM Tris-HCI buffer on DEAE Sephadex column, principal lectin was identified.

• Journal of Microbiology and Biotechnology
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• v.14 no.5
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• pp.891-900
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• 2004

UDP-N-Acetylglucosamine(GlcNAc):$\beta$1,4-D-mannoside$\beta$-l ,4N-acetylglucosaminyltransferase-III (GnT-III) and UDP-N-GlcNAc:$\alpha$-6-D-mannosid$\beta$-1,6N-acetylglucosaminyltransferase-V(GnT - V) activities were determined in human hepatoma cell lines and metastatic colon cancer cells, and their activities were compared with those of normal liver cells and fetal hepatocytes. GnT-III activities were higher than those of GnT-V in hepatic carcinoma cells. When the two enzyme activities were assayed in highly metastatic colon cancer cells, GnT - V activities were much higher than those of GnT-III. When GlcN, GlcN-biant-PA and UDP-GlcNAc were used as substrates, the enzymes displayed different kinetic properties between hepatic and colon cancer cells, depending on their metastatic potentials. Normal cells of two origins had characteristically very low levels of GnT-III and -V activities, whereas hepatoma and colon cancer cells contained high levels of activities. These data were supported by RT-PCR and Northern blot analyses, showing that the expression of GnT-III and -V mRNAs were increased in proportion to the enzymatic activities. The increased GnT-III, md -V activities were also correlated with increased glycosylation of the cellular glycoproteins in hepatoma and colon cancer cells, as examined by lectin blotting analysis by using wheat germ glutinin (WGA), erythroagglutinating phytohemagglutinin (E-PHA), leukoagglutinating phytohemagglutinin (L-PHA), and concanavalin A (Con A). Treatment with retinoic acid, a differentiation agent, resulted in decreases of both GnT-III and -V activities of HepG2 and HepG3 cells. In colon carcinoma cells, however, treatment with retinoic acid resulted in a reduction of GnT-V activity, but not with GnT-III activity. Although the mechanism underlying the induction of these mzymes is unclear, oligosaccharides in many glycoproteins have been observed of cancer cells.

• Current Research on Agriculture and Life Sciences
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• v.7
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• pp.83-97
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• 1989

This study was carried out to obtain basic information necessary for aggregation and in-vitro culture of mouse embryos by treating phytohemagglutinin-p (PHA-P). The 4-, 8-cell and morula embryos were obtained from female mice of albino BALE/C, CBA and C57BL strains, those were injected 5 i.u pregenant mare serum gonadotrophin and 5 i.u human chorionic gonadotrophin to superovulation. The zona pellucidia was removed by placing the embryos in Acidic Tyrode solution containing 1.0% protease or/and 5 ug/ml PHA-P. The pairs of zona free embryos were subjected to aggregation by glassneedle in BMOC-3 containing 5 ug/ml PHA-P. The aggregation embryos were cultured in Brinster's mouse ova culture-3(BMOC-3) medium under the gas phase of 5% $CO_2$ in air $37^{\circ}C$ for 13 to 50 hours. The results obtained in this study are summarised as follows : 1. When 4-, 8-cell and morula embryos were zona-freed in acidic Tyrode solution containing 1.0% protease or/and 5 ug/ml PHA-P, and cultured in vitro to blastocysts, the 4- and 8-cell embryos showed slightly less development rates than the morula one did, and solution of 5 ug/ml PHA-P brought some higher development rate than negative control. 2. As 2, 5 or 10 ug/ml PHA-P was added to the solution to aggregate 4-, 8-cell or morula embryos, 2 ug/ml solution represented slightly lower aggregation rate than the higher levels solutions, and 4- and 8-cell embryos showed higher rates than morula one did (P<.05). 3. In respect to the development rates of aggregated embryos to morula no significant difference was found among PHA-P levels and between 4-and 8-cell embryos. With respect to those of aggregated embryos to blastocysts the different levels of PHA-P showed similar results, however, the 4- and 8-cell embryos represented higher rates than the morula one did (P<.05). 4. The mean time necessary for development of aggregated 4-, 8-cell and morula embryos to blastocysts were 38.5-40, 26-27 and 19-20hrs. Respectively in solution for aggregation. 5. The aggregation rates of embryos were 34-94%, when treated protease or/and PHA-P. Supplementation of 5 ug/ml PHA-P to the solution for aggregation showed a trend demonstrating higher aggregation rate compared to negative control, although no significance was found. However, 4- and 8-cell embryos represented significantly higher aggregation rates than the morula one did (P<.05). 6. The development rates of 4- and 8-cell embryos to morula were 52.7-84.7 and 73.8-87.2%, respectively, showing no significant difference between two cell stages. However, the aggregation rates of embryos treated with solution containing PHA-P were higher than negative control (P<.05). 7. The development rates of 4- and 8-cell and morula embryos to blastocysts were 41.7-77.7 78.7-83.0 and 0-19.2%, respectively. The rates of 4-cell embryos treated with PHA-P were significant higher than the negative control (P<.05). The 8-cell and morula embryos also showed more rates when treated PHA-P.

• Journal of Embryo Transfer
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• v.9 no.1
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• pp.111-116
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• 1994

This study was carried out to investigate the aggregation rate of isolated mouse 2-, 4- and 8-cell stage blastomeres in phytohemagglutinin(PHA) solution. The morphologically normal embryos were collected from the oviduct of superovulated female mouse by flushing with M2 and the zona pellucida of embryos were removed with 0.5% pronase. The blastomeres were isolated by pipetting after plunging into Ca++-Mg++free PBS for 20 min. The result showed that aggregation rate in 0.5% (84.9~93.1%) was higher than that in 1.0% PHA(76.0~82.1%). Optimal aggregation time was 60min (83.9~100.0%) when compared with 30min (78.8~87.5%). Developmental to blastocyst in recombinated blastomeres was higher under conditions of 0.5% PHA solution and 60-min aggregation than that under other conditions.

• Korean Journal of Veterinary Research
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• v.34 no.2
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• pp.301-306
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• 1994

This study was planned to estimate the activity of bovine circulating blood lymphocytes using phytohemagglutinin-M(PHA) known as T cell mitogen. Bovine circulating blood mononuclear cells(MNCs) was separated, and cultured with or without macrophage($PHA^+/M{\phi}^+$ or $PHA^+/M{\phi}^-$) in conditioned medium which stimulated with various concentration of PHA(0, 5, 10, 15 and $20{\mu}g/ml$ in medium), and then investigated the blastogenic response and rosette formation of lymphocytes. Blastogenic rate(BR) was especially increased in PHA concentration(10 and $15{\mu}g/ml$) of $PHA^+/M{\phi}^+$ group and their BR were $41.5{\pm}6.8%$ and $44.4{\pm}8.9%$, respectively and BR in PHA concentration(15 and $20{\mu}g/ml$) of $PHA^+/M{\phi}^-$ group was $32.8{\pm}6.2%$ and $31.4{\pm}4.6%$, respectively. BR of lymphocytes was more increased in $PHA^+/M{\phi}^+$ than $PHA^+/M{\phi}^-$ group when these cells were stimulated by PHA. Rosette forming rate(RFR) of lymphocytes to SRBC highly increased when SRBC was treated with AET and/or dextran, respectively. On the orther hand, RFR significantly increased more in $PHA^+/M{\phi}^+$ and $PHA^+/M{\phi}^-$ group than in control group, but when compared with two groups, statistical significancy was recognized only in PHA concentration($15{\mu}g/ml$, p<0.026) of $PHA^+/M{\phi}^+$ group.

• Journal of Nutrition and Health
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• v.21 no.2
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• pp.113-121
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• 1988

The purpose of this study was to investigate the effects of dietary zinc on immune response in mice. Weanling male mice was placed individually in stainless steel cages and fed a zinc dificient diet and control diet. All mice were given deionized water ad libitum. The introduction of extraneous zinc was minimized in all cage by washing feed jars and water bottles sequentially with 4mM EDTA and conc-nitiric acid followed by deionized water. After 4 and 5 weeks of the diets, mice were immunized with lx 106 Naegleria fowleri intraperitoneally. Mice were weighed once a week. The results from this study are summarized as followed ; 1) Mice fed the zinc dificient diet showed growth retardation. After 3 weeks of diets, mean body weight of zinc deficient mice was 21.4g and that of control was 25.0g. This difference is singnificant statistically (p<0.01). The more time passed, the more remarkable difference was found. 2) The weigth of organs were measured on liver, kidney, spleen, thymus, heart, lung, brain. Difference in weight were observed only in liver and spleen. 3) Proliferative response of spleen cells of zinc deficient mice to con A was lower than that of control mice after one week on immunization(p<0.005). 4) Stimulation index was lower in zinc deficient mice to phytohemagglutinin after two weeks on immunization (p<0.05). 5) Blastogenesis of speen cells of zinc deficient mice to Naegleria fowleric lysate was lower after 10 days on immunization (p<0.05). 6) Immunoglobulin G antribody titers of zinc deficient mice sera by ELISA was lowered to control mice after 5 weeks on immunization (p<0.005).

• Journal of the Korean Society of Food Science and Nutrition
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• v.26 no.6
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• pp.1194-1199
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• 1997

The effect of hot taste preference on selected immune responses was investigated in human peripheral immunocompetent cells. Human lymphocytes and natural killer(NK) cells were prepared at a concentration of 2$\times$$10^{6}$ cells/ml in RPMI-1640 containing 10% fetal bovine serum. Lymphocytes proliferation was determined with the [$^{3}H$]-thymidine pulse for 18hrs after concanavalin A, phytohemagglutinin, Salmonella typhimurium mitogen, or media alone. NK cell activity was measured by cytolysis of $^51Cr$-labeled target cells K562. Serum antibodies levels such as IgM, IgG, IgA were also measured by ELISA method. There was no difference of serum IgM level among the groups, but IgG and IgA levels were greater in the group with hot taste preference than those of the group without hot taste preference. In lymphocytes of the group with hot taste preference there was a greater mitogen-induced lymphocyte proliferative responses compared to the group without hot taste preference. In addition, NK cell activity in group with hot taste preference was lower than that of the group without hot taste preference. These results suggest that the eating habit of spicy food containing hot components may affect immune status by modulating selective immunocompetent cells function.

• Asian-Australasian Journal of Animal Sciences
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• v.25 no.7
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• pp.1038-1044
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• 2012

In the present study, the impact of Salmonella Typhimurium on cell-mediated immunity (CMI) was investigated in 5 week-old immuno divergent broiler lines selected for the high and low response to phytohemagglutinin-P. The immune response was assessed in peripheral-blood mononuclear cells (PBMCs) induced with Salmonella Typhimurium at different time intervals (0 h, 0.5 h, 2 h, 4 h, 6 h, 12 h and 24 h). The differential mRNA expression patterns of IFN-${\gamma}$, IL-2 and iNOS were evaluated by quantitative real time PCR. In-vitro production of nitric oxide (NO) was also estimated in the culture supernatant and correlated with iNOS mRNA expression. Present study showed higher production of NO in the high cell-mediated line (HCMI) as compared to the low cell-mediated line (LCMI) upon stimulation with Salmonella Typhimurium. Correspondingly, higher mRNA expression of iNOS and IFN-${\gamma}$ were observed in high response birds (HCMI); but IL-2 was down regulated in this line compared to the low response birds (LCMI). Significantly (p<0.05) higher expression of iNOS, IFN-${\gamma}$ and higher production of NO in high line indicated that the selection for PHA-P response might be employed for increasing the immune competence against Salmonella Typhimurium in chicken flocks.