Removal of ligand-bound liposomes from cell surfaces by microbubbles exposed to ultrasound

Gas-filled microbubbles attached to cell surfaces can interact with focused ultrasound to create microstreaming of nearby fluid. We directly observed the ultrasound/microbubble interaction and documented that under certain conditions fluorescent particles that were attached to the surface of live cells could be removed. Fluorescently labeled liposomes that were larger than 500 nm in diameter were attached to the surface of endothelial cells using cRGD targeting to αvβ3 integrin. Microbubbles were attached to the surface of the cells through electrostatic interactions. Images taken before and after the ultrasound exposure were compared to document the effects on the liposomes. When exposed to ultrasound with peak negative pressure of 0.8 MPa, single microbubbles and groups of isolated microbubbles were observed to remove targeted liposomes from the cell surface. Liposomes were removed from a region on the cell surface that averaged 33.1 μm in diameter. The maximum distance between a single microbubble and a detached liposome was 34.5 μm. Single microbubbles were shown to be able to remove liposomes from over half the surface of a cell. The distance over which liposomes were removed was significantly dependent on the resting diameter of the microbubble. Clusters of adjoining microbubbles were not seen to remove liposomes. These observations demonstrate that the fluid shear forces generated by the ultrasound/microbubble interaction can remove liposomes from the surfaces of cells over distances that are greater than the diameter of the microbubble.     

Onset of puberty in CD-1 mouse pups exposed prenatally through weaning to endophyte-infected tall fescue seed

This study was undertaken to investigate the effects of feeding endophyte - infected (Acremonium coenophialum ) tall fescue seed to CD-1 mouse dams (P(1)) during gestation and lactation, and on the subsequent growth and sexual maturity (onset of puberty) of their male and female offspring (F(1)). Forty-eight 21 d old pups (24 male and 24 female F(1) mice) were weaned from dams fed one of two diets containing 50% rodent chow (w/w) and 50% KY-31 tall fescue (Festuca arundinacea ) seed. The seed in Diet 1 was noninfected, while the seed in Diet 2 was 80% endophyte-infected. At weaning (21 d), the F(1) pups were fed rodent chow, ad libitum throughout the remaining experimental period. At 24 d, they were paired with sexually mature non-treated virgin CD-1 mice (fed 100% rodent chow) for one parturition cycle. Male F(1) mice were sacrificed at 84 d to determine testicular development. The age at the birth of the first litter for Diet 2 F(1) male (76.8 +/- 2.2 d) and female (58.4 +/- 2.1 d) was significantly greater (P<0.05) than the age at parturition for Diet 1 male and female F(1) test mice (64.1 +/- 1.8 and 51.9 +/- 1.2 d, respectively). At parturition, the female F(1) mice showed no significant differences (P>0.05) in either mean parturition weight or number of F(2) pups born per litter. However, total F(2) litter wight (11.38 +/- 1.14 g) and mean weight per F(2) pup (1.40 +/- 0.04 g) for Diet 2 female F(1) mice litters were lower (P<0.05) when compared with Diet 1 females (14.53 +/- 0.57 g and 1.66 +/- 0.02 g, respectively). No significant differences were observed between the two male F(1) treatment groups, for total F(2) litter weight or the number of pups born per F(2) litter. Although Diet 2 F(1) males weighed significantly less (P<0.05) at weaning and at pairing, final body weights at sacrifice (84 d) were not different (P>0.05) from the Diet 1 males.     

Suckling deficits in rat pups exposed to alcohol in utero

With the aid of a pair-feeding procedure, two groups of pregnant Long-Evans rats were fed a liquid diet containing 35% or 0% ethanol-derived calories during days 6-20 of gestation. A third group was allowed free access to standard lab chow and water throughout pregnancy. At 6-7 or 9-10 days of age, suckling performance by male and female offspring representing the three prenatal treatment groups was examined. The test stimulus was a 6-10-day postparturient, anesthetized dam in which milk letdown was prevented. Compared to both pair-fed and lab chow controls, alcohol-exposed animals exerted a lower maximum suckling pressure, spent less time suckling during the test session, and displayed an altered suckling pattern. These data are consistent with existing clinical and experimental evidence documenting sucking deficits following prenatal alcohol exposure and are discussed in terms of prenatal alcohol-induced CNS impairment.     

Production of mouse pups from germline transmission-failed knockout chimeras

Occasionally, chimeras do not transmit the gene of interest to pups in gene disruption experiments. However, the risk of failure could be reduced if we could identify embryonic stem (ES)-derived germ cells in the testis. Here, we report the production of pups from three lines of infertile chimeric male mice and the establishment of knockout lines by combining green fluorescent protein-tagged ES cells with intracytoplasmic sperm injection.     

The ultrasonic responses of albino mouse pups to tactile stimuli

Tactile stimuli, like environmental temperature changes, can evoke ultrasonic responses from albino mouse pups. But the changes with age in the intensity of ultrasounds so produced follow a different pattern from those due to temperature changes. The responses begin with very high intensity pulses in the very young pups and then gradually decline with the age of the pups. The present report arises from a systematic study of this phenomenon, the results of which are discussed in relation to those of previous ones.     

Induction of maternal nest building in virgin female mice by the presentation of young

Ovariectomized and intact adult virgin female mice presented with two 1-day-old mouse pups built larger nests and more nests rated “maternal” than did intact animals not proffered young. Virgin females presented with live 1-day olds behind a wire partition in their homecages also constructed larger nests and more nests rated “maternal” as compared to animals presented with freshly killed 1-day olds, 19-day olds, or with no pups behind the partition.     

Survival of cultured mammalian cells exposed to ultrasound

Summary The colony-forming ability of cultured mammalian cells exposed to monochromatic ultrasonic vibrations of different frequencies (0.1, 0.5, 1.0, 2.0, 3.3 MHz) was studied. The combined effect of x rays and 1.0-MHz ultrasonic waves on the survival of M3-1 cells was also investigated. Split-dose experiments showed that cells exposed to ultrasound are sensitized to a subsequent exposure. Almost twice as many cells survive a given ultrasonic dose when exposed in the M phase as when exposed in the S phase. A small amount of synergism between ultrasound and x rays has been observed. The extent of synergism depends on the experimental factors, and may be due to an interaction between nuclear damage caused by x rays and the damage to the cell membrane caused by ultrasound.Research supported by the U.S. Atomic Energy Commission and the National Aeronautics and Space Administration     

Effects of ultrasound on the mouse exposed at different stages of gestation: Acute studies

Summary At five stages of gestation, mice were exposed dorsally to ultrasonic irradiation at 1 W/cm² (spatial and temporal average), 2 MHz, and 34 and 37° C (temperature at beginning of exposure). The results indicated that ultrasonic exposure up to at least 100 s is not hazardous to a pregnant mouse or its offspring in utero under the conditions of our experiment. Thresholds for both undesirable and lethal effects on the dam and its embryos or fetuses appeared between 100 and 200 s. Temperature rises in the uterus to more than 40° C and similarity of results obtained with continuous-wave and burst modes of irradiation suggest a thermal mechanism as one cause of these effects. Preliminary conclusions are that thresholds for both tissue damage to the dam and deleterious effects on pregnancy were similar; however, they occurred at spatial average intensity and exposure levels far in excess of those used in clinical diagnostic medicine.     

Chromosomic aberrations in female workers exposed to pesticides

The purpose of this work was to determine if the occupational exposure to those pesticides used at banana plantations' packaging plants produces genetic damage to somatic cells of female workers. Chromosomal aberrations were scored in lymphocytes of 20 women, 10 female exposed workers and 10 female controls. Workers were recruited from independent farms from two locations in Costa Rica, during January through June in 1996 and 1997. These females had a minimum of three months of work, had never received chemotherapy or radiotherapy and did some of these labors: sealing, spraying or weighting of bananas. Control unexposed females lived in the same area, were of similar age and neither them nor their husbands/mates had ever worked in pesticide related labors. For each female, 100 mitotic figures were scored. The kind of aberrations detected were acentric fragments, dicentric chromosomes, rings, gaps and breaks. Among workers, 16% of cells (n=1000) had one or more abnormalities, whereas control unexposed females had 6% of cells (n=1000) with comparable anomalies (p < 0.05). In conclusion, the pesticide exposure is a risk factor for chromosome aberrations in female somatic cells.     

Pregnancy in female house mice exposed to urinary chemosignals from other females.

Seven experiments were performed to investigate pregnancy termination, urinary chemosignals, and litter sex ratio variation in female house mice. Experiments tested the effects of urine from adult and prepubertal females, housed individually or in groups, on successful insemination and litter production by females treated at different times and for different periods during the 3 weeks before mating and during gestation. Treatment of females with urine from adult females housed eight per cage or with urine pooled from eight adult females housed individually for 2 or 3 weeks before mating resulted in fewer successful pregnancies and significantly more female-biased litters. Treatment with urine from adult or prepubertal females housed eight per cage or with urine pooled from eight mice housed individually for the first 6 days of gestation or throughout pregnancy resulted in a significant increase in the rate of pregnancy termination. These treatments resulted in lower body weights at birth and slower growth rates in all males and in some females. Puberty was delayed in female progeny from urine-treated dams in five of seven experiments, and these young females attained first oestrus at greater mean body weights than mice in other treatments. These findings indicate that, in mice, at high population density, communication via a urinary chemosignal can alter reproduction in recipient females. Availability of, and competition for, resources such as food would be greater at higher densities, possibly lowering the probability of reproductive success. Pregnancy termination and delays in reproduction and attainment of sexual maturity might lead to greater successful reproduction at a later time.     

Hatching of oothecae from female Blattella germanica exposed to hydramethylnon and boric acid baits

Adult female Blattella germanica (L.) (Dictyoptera: Blattellidae) carrying oothecae were confined in cages with bait formulations of boric acid or hydramethylnon at 90% r.h. and either 20° or 28° C until all had been killed. Females poisoned by hydramethylnon retained their oothecae at death, while a considerable proportion of those killed by boric acid deposited their oothecae prematurely. Oothecae from both groups of insects were monitored until they either hatched or lost viability. Oothecae from both 7–14 day old adults and 14–21 day old adults poisoned with hydramethylnon showed reduced hatching relative to controls at 20° but not at 28 °C, an effect attributed to premature loss of water-exchange between oothecae and female adult during the maturation process. Fewer oothecae hatched from females poisoned with boric acid at both temperatures, and the proportion of those which did hatch was higher in the group retained by the female at death than in the group prematurely deposited. These results for the boric acid-treated females and results which showed no significant difference between the hatch of oothecae from hydramethylnon-treated females and decapitated females suggested that transfer of insecticide from treated female to oothecae was not important in reducing hatch for either compound. Reinfestation as a consequence of the failure of conventional blatticides to prevent hatching of oothecae carried by female B. germanica is discussed.

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Résumé Des femelles adultes de B. germanica, maintenues à 20° ou 28° C et 90% HR, ont été exposées, 7 à 14 jours ou 14 à 21 jours après l'émergence à des appâts contenant de l'hydraméthylnone ou de l'acide borique. Empoisonnées par l'hydraméthylnone les femelles retiennent leur oothèque jusqu'à la mort, tandis que empoisonnées par l'acide borique, elles déposent leur oothèque prématurément. Toutes les oothèque des femelles empoisonnées ont été maintenues dans les mêmes conditions écologiques jusqu'à ce qu'elles éclosent ou se révèlent non viables. Avec l'acide borique, une réduction des naissances a été observée pour les oothèques des mères des 2 classes d'âges et aux 2 températures, ce qui n'a eu lieu qu'à la température basse avec l'hydraméthylnone. L'effet ne semble pas dû au transfert de l'insecticide de la femelle à l'oothèque et est plus probablement provoqué par la fin des échanges d'eau entre la mère et l'oothèque, à la mort ou lors de l'émission prématurée. Pour les femelles traitées à l'acide borique, la proportion d'éclosion d'oothèques est plus importante chez les femelles ayant retenu leur oothèque jusqu'à la mort que chez celles qui les ont émises prématurément. Les femelles empoisonnées lors d'opérations d'éradication peuvent mourir ou déposer leurs oothèques loin des zones traitées; les larves éclosant de ces oothèques peuvent maintenir la contamination.

     

Reproductive aspects in female rats exposed prenatally to hydrocortisone

We investigated the effects of hydrocortisone during the prenatal period and its later repercussion on reproductive aspects of female rats. Pregnant rats were treated (s.c.) with hydrocortisone acetate, at 1.5 mg/day on the 17th, 18th, and 19th days of pregnancy. Although the present study was not intended to identify mechanisms of toxicity, the treatment with hydrocortisone in the last period of pregnancy presented no signs of toxicity. The efficacy of the hydrocortisone in reducing the adrenal wet mass and plasma corticosterone levels immediately after delivery in both the treated mothers and in respective pups at birth may indicate impairment of the hypothalamus-pituitary-adrenal axis. In addition, the treatment with hydrocortisone did not interfere in the development of the female descendants until puberty. However, it affected the estrous cycle and fertility. Probably, the prenatal exposure to corticosteroids had altered at least partially the hypothalamus-pituitary-gonadal axis, resulting in the damages observed in adult life. These results indicate that the use of the hydrocortisone at a dose that apparently does not endanger the neonate led to undesirable effects in the adult reproductive phase, resulting in later deleterious alteration of the reproductive physiology in female rats.     

Sensory processes in the discrimination of pups by female mice (Mus musculus)

Maternal pup retrieval by house mice (Mus musculus) was studied in an investigation of sensory processes in the discrimination of ‘own’ from ‘alien’ pups. In a control condition, mothers were able to discriminate their own from alien young. This ability seemed to be based on olfactory cues, while visual and auditory cues did not seem to be involved. Furthermore, it is suggested that retrieval consists of: elicitation (probably mediated by auditory cues); location of the pup (auditory and olfactory cues); and recognition of the pup (initially olfactory cues, with subsequent gustatory and tactile cues).     

Development of the central nervous system functions in rat pups prenatally exposed to alcohol (study on the behavioural teratology of ethanol in CFY rat pups)

As a model of fetal alcohol syndrome (FAS) the rate of the maturation of the functions of the central nervous system was studied in the offspring of pregnant CFY rats receiving (from the 7th-15th day of gestation) either oral ethanol treatment or liquid diet containing ethanol. Both types of exposure caused numerous behavioural impairments, besides high perinatal mortality also the opening of the eyes and ears, and the appearance of postural reflexes were delayed. The newborn rats could be characterized by hyperactivity and weak motor coordination. The learning capacity, the avoidance conditioned reflexes was the poorest in the case of the offspring of mothers kept on liquid diet, containing alcohol, the latency of the conditioned response was significantly lenghtened. During reconditioning, in the case of the sexually already mature pups, the weakest performance was observed in the offspring of mothers having received oral alcohol treatment. This findings indicated, on one hand, that the retardation ceased and, on the other, that the learning and memory impairments caused by oral alcohol exposure was persistent. Following prenatal alcohol exposure carried out by different methods the neurotoxic effect, the retardation of the rate of maturation of the central nervous functions, and the adaptive mechanisms were all affected to different extent. Besides alcohol exposure also other factors (relative protein insufficiency, malnutrition) may be involved in the pathomechanism of the above mentioned phenomena.     

Gonadal hormones stimulate ultrasound production by female hamsters.

Two studies were conducted to examine the role of sex hormones in ultrasound production by female hamsters. Ovariectomized hamsters were treated with estradiol benzoate (EB), testosterone propionate (TP), progesterone (P), estradiol plus progesterone (EB + P), testosterone plus progesterone (TP+P), or oil vehicle. Rates of ultrasound production by these females were observed in response to brief male-female contact, and during exposure to synthetic ultrasounds. Maximal rates of ultrasound production required EB + P or TP+P replacement therapy. Intermediate call rates were stimulated by EB and TP individually, whereas P alone had no significant effect. These results support the hypothesis that ovarian estrogens and progesterone control proceptive as well as receptive behaviors in female hamsters.