Full-term development of mouse blastomere nuclei transplanted into enucleated two-cell embryos

The nuclei from four- and eight-cell mouse embryos were transplanted into enucleated two-cell embryos. It was found that such embryos not only developed to the blastocyst stage in vitro (72% and 35%), but also developed to full term (22% and 8%) after transfer to recipient mice. However, development of embryos which contained nuclei from the inner cell mass was not observed. Since the development of enucleated zygotes which contain advanced nuclei is limited (the present study; McGrath and Solter: Science, 226:1317-1319, '84; Robl, Gilligan, Critser, and First: Biol. Reprod., 34:733-739, '86), it appears that cytoplasmic factors are important for the development of nuclei from advanced cells.     

Stimulation of the interaction between actin and myosin by Physarum caldesmon-like protein and smooth muscle caldesmon.

We have purified an actin-binding protein from the plasmodia of a lower eukaryote, Physarum polycephalum, with an apparent molecular mass of 210,000 daltons on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This protein bound to actin filaments with a stoichiometry of 1:7-8 in a Ca(2+)-calmodulin-dependent manner. Antibody raised against caldesmon from smooth muscle cross-reacted with the 210-kDa protein. In vitro motility assay revealed that the 210-kDa protein increased the sliding velocity of actin filaments on Physarum myosin. The 210-kDa protein more than doubled the actin-activated ATPase activity of Physarum myosin under comparative conditions of in vitro motility assay. Further increases in the concentration of the 210-kDa protein decreased its stimulatory effects. Ca(2+)-calmodulin prevented the stimulatory effects of the 210-kDa protein. Unexpectedly, smooth muscle caldesmon also increased the sliding velocity of actin filaments on smooth muscle myosin at lower concentrations. The well-known inhibitory effect of smooth muscle caldesmon on the actin-myosin interaction was observed with this motility assay when the concentration of the caldesmon was increased further. The stimulatory and inhibitory effects were confirmed by measurements of actin-activated ATPase activity of smooth muscle myosin. From estimations of the intracellular concentrations of the 210-kDa protein and smooth muscle caldesmon in vivo, it appears that effects of the former and the latter on actin-myosin interactions in vivo are stimulatory and inhibitory, respectively.     

A computational aspect of kinetic depth effect

A method is proposed to determine the rigid structure as well as the three dimensional motion of an object from a sequence of orthographically projected images. It is assumed that the velocities as well as the positions of the points attached to the object are observable in the images. The instantaneous rigidity condition wich states that the relative position vector between any two points is orthogonal to the corresponding relative velocity vector is derived from the condition of rigidity. Assuming further that the rotational velocity component is constant throughout the period of observation, each of the projected relative velocity vectors is shown to move along one of the elliptical trajectories, all of which are similar to one another. The orientations of the longer axes are common to every trajectory. Then one can determine the equations of the ellipses by observing only two points in three views or three points in two views. The solution is obtained from a set of linear equations. The length of the longer axis enables one to determine the relative velocity. The instantaneous rigidity condition is then used to obtain the relative position. The special cases where the rotational axis is either perpendicular to or parallel with the image plane are discussed. The above results are discussed in relation to the relevant psychophysical observations as well as theoretical studies.     

Quantitative determination of titers of anti-zona serum.

The titers of rabbit antiserum against isolated mouse zonae pellucidae were examined by several methods in connection with inhibitory effect on fertilization. The titers determined by zona precipitate, zona dissolution, indirect immunofluorescence and in vitro fertilization tests were 2(4), 2(1) to 2(4), 2(7) and 2(4), respectively. Cytotoxic effect could not be detected from zona antibody. The indirect immunofluorescence was most sensitive for detection of zona antibody but did not represent the extent of inhibitory effect on fertilization. The titer obtained by zona precipitate test was most close to the titer obtained by inhibitory effect on in vitro fertilization. The present study also demonstrated that at least 0.0375 ml of antiserum per female mouse, equivalent to 0.15-0.25% of body weight, was necessary for inhibition of fertilization in vivo by passive immunization with anti-zona serum.     

A new approach for mealybug management: recruiting an indigenous,but ‘non‐natural’ enemy for biological control using an attractant

We previously discovered that (2,4,4‐trimethyl‐2‐cyclohexenyl)‐methyl butyrate (cyclolavandulyl butyrate, CLB) is an attractant for the mealybug‐parasitic wasp Anagyrus sawadai Ishii (Hymenoptera: Encyrtidae: Anagyrini). This wasp is not likely to parasitize the Japanese mealybug, Planococcus kraunhiae (Kuwana) (Hemiptera: Pseudococcidae), under natural conditions. In this study, we showed that this ‘non‐natural’ enemy wasp can parasitize P. kraunhiae in the presence of CLB in field experiments. Laboratory‐reared mealybugs placed on persimmon trees with CLB‐impregnated rubber septa were parasitized significantly more often by endoparasitic wasps than those on non‐treatment trees (18.1–40.3 vs. 0–6.3%). Anagyrus sawadai accounted for 20% of the wasps that emerged from mealybugs placed on CLB‐treated trees. Moreover, CLB attracted another minor parasitoid, Leptomastix dactylopii Howard (Hymenoptera: Encyrtidae: Anagyrini), which also parasitized more P. kraunhiae in the presence of CLB. All wasps that emerged from the mealybugs on control trees were Anagyrus fujikona Tachikawa, a major parasitoid of P. kraunhiae around the test location. These results demonstrated that CLB can recruit an indigenous, but ‘non‐natural’ enemy that does not typically attack P. kraunhiae under natural conditions, as well as a minor natural enemy, for biological control of this mealybug species.     

Overexpression of wheat alternative oxidase gene Waox1a alters respiration capacity and response to reactive oxygen species under low temperature in transgenic Arabidopsis

Under low temperature conditions, the cytochrome pathway of respiration is repressed and reactive oxygen species (ROS) are produced in plants. Mitochondrial alternative oxidase (AOX) is the terminal oxidase responsible for the cyanide-insensitive and salicylhydroxamic acid-sensitive respiration. To study functions of wheat AOX genes under low temperature, we produced transgenic Arabidopsis by introducing Waox1a expressed under control of the cauliflower mosaic virus (CaMV) 35S promoter in Arabidopsis thaliana. The enhancement of endogenous AOX1a expression via low temperature stress was delayed in the transgenic Arabidopsis. Recovery of the total respiration activity under low temperature occurred more rapidly in the transgenic plants than in the wild-type plants due to a constitutively increased alternative pathway capacity. Levels of ROS decreased in the transgenic plants under low temperature stress. These results support the hypothesis that AOX alleviates oxidative stress when the cytochrome pathway of respiration is inhibited under abiotic stress conditions.     

The induction of murine tumor infiltrating lymphocytes (TIL) by interleukin-2 or T cell growth factor

Mice were injected in the foot pad with either 5×10⁵ syngeneic plasmacytoma (MOPC104E) or fibrosarcoma cells (Meth A). Lymph nodes containing tumor cells were harvested 14 days later and cultured. In the presence of recombinant interleukin-2 (r-IL-2) predominantly tumor cells proliferated. Culture with T cell growth factor (TCGF) resulted in the growth of lymphoid cells. Concanavalin A (Con A) had only a modest effect on elimination of tumor cells in the culture. Tumor-infiltrating lymphocytes (TIL) prepared from the lymph nodes showed specific tumor-neutralizing activity when grown in the presence of TCGF. In vitro examination revealed that Meth A cells could not be lysed by TIL, while TIL from MOPC tumors showed tumor specific activity. This study may explain negative results in human trials with TIL induced by IL-2 alone.Abbreviations r recombinant - IL-2 interleukin-2 - TCGF T cell growth factor - TIL tumor infiltrating lymphocytes - Con A concanavalin A - HBSS Hanks' balanced salt solution     

Sequential immunotherapy using interleukin-1 followed by interleukin-2 of ascitic MOPC104E-bearing mice

This study shows that intraperitoneal injection of interleukin-1 (IL-1), followed by interleukin-2 (IL-2), can effectively eradicate murine ascitic tumor cells. This antitumor effect of IL-1 and IL-2 was abolished when administration of IL-2 preceded that of IL-1. Solid tumors inoculated subcutaneously (s.c.) into the back of mice were also sensitive to this combined i.p. therapy, indicating a systemically-operating antitumor mechanism. Splenocytes from tumor-bearing mice treated with IL-1 followed by IL-2 showed a strong tumor-neutralizing activity. The population responsible proved to be Lyt2.2 (CD8)-positive cells.Abbreviations IL interleukin - LAK lymphokine activated killer - LU lytic unit - MST median survival time - SE sonicated tumor extract