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Focuses on the provision of child protection services (CPS) in the U.S. Benefits of CPS program; Creation of the Child and Family Service Review process; Policy guidelines of child abuse.

X-linked severe combined immunodeficiency (XSCID) is a life-threatening syndrome in which both cellular and humoral immunity are profoundly compromised. This disease results from mutations in theIL2RG gene, which encodes the common cytokine receptor ? chain, ?c. Previously, we generated ?c-deficient mice as a murine model of XSCID. We have now used lethally irradiated ?c-deficient mice to evaluate a gene therapeutic approach for treatment of this disease. Transfer of the human ?c gene to repopulating hematopoietic stem cells using an ecotropic retrovirus resulted in an increase in T cells, B cells, natural killer (NK) cells, and intestinal intraepithelial lymphocytes, as well as normalization of the CD4:CD8 T-cell ratio and of serum Ig levels. In addition, the restored cells could proliferate in response to interleukin-2 (IL-2). Thus, our results provide added support that gene therapy is a feasible therapeutic strategy for XSCID. Moreover, because we used a vector directing expression of human ?c to correct a defect in ?c-deficient mice, these data also indicate that human ?c can cooperate with the distinctive cytokine receptor chains such as IL-2Rß and IL-7R? to mediate responses to murine cytokines in vivo.

The ecdysone titer of larvae and pupae of Chironomus thummi was determined by radioimmune assay (RIA) and revealed a concentration of about 150 ng/g fresh weigth in late third instar larvae and a peak of more than 450 ng/g fresh weight just preceding pupation. The ecdysone titer curve shows a high degree of correlation with previously observed puffing activity at the ecdysone sensitive chromosomal site IIIdl. Further, β-ecdysone is the only endogenous ecdysone detected by high pressure liquid chromatography during larval-pupal development. It was also observed that developmentally arrested Chironomus contain less ecdysone than “normal” larvae.

1.A 28-kDa peptide from the brain of the tobacco hornworm,Manduca sexta, was purifiedvia HPLC. The peptide copurified with the insect neurohormone, prothoracicotropic hormone (PTTH), through two HPLC columns.2.Immunocyctochemistry using polyclonal antibodies against the 28-kDa peptide revealed that the peptide was produced in the same protocerebral neurons that produce PTTH. Western blot analysis demonstrated that the 28-kDa peptide and big PTTH are different molecules.3.A PTTHin vitro bioassay indicated that despite having chromatographic properties similar to those of big PTTH and being produced by the same neurons, the 28-kDa peptide did not have PTTH activity.4.Amino acid sequence analysis yielded a 27 N-terminal amino acid sequence that had no similarity with known peptides.5.Immunocytochemical studies revealed that the 28-kDa peptide is present as early as 30% embryonic development and is absent by adult eclosion. This is in contrast to big PTTH, which is expressed throughout theManduca life cycle.6.These data suggest that the 28-kDa peptide is another secretory phenotype of the lateral neurosecretory cell group III (L-NSC III) which may have functions distinct from those for big PTTH or may act synergistically with big PTTH.

Head segments and brains were extirpated from embryos of the tobacco hornworm,Manduca sexta, extracted and the resulting extracts assayed for prothoracicotropic hormone (PTTH) activity on prothoracic glands from day 3 fifth instar larvae and day 0 pupae. Dose-response curves were generated and indicated the presence of PTTH activity in embryonic brains and head segments, suggesting a role(s) for this neurohormone during embryogenesis. Maximal PTTH activity was found in brains from embryos 117 h post-oviposition, just prior to hatching, but activity was also noted in head segments as early as 24 h postoviposition. These data on PTTH and those on ecdysteroids and juvenile hormones in embryos suggest that these 3 classes of hormones which control insect post-embryonic development, may also be involved in the regulation of developmental processes in the embryo.

We have previously shown that basic fibroblast growth factor (bFGF) is mitogenic for human bone marrow stromal cells and enhances myelopoiesis in human long-term bone marrow culture. In the present study, we examined the mechanism by which bFGF enhances granulopoiesis. We observed that bFGF significantly abrogated the inhibitory effect of transforming growth factor-β1 (TGF-β1) on granulocyte-macrophage colony-stimulating factor (GM-CSF)-supported progenitor cell growth (P= .009). The partial reversal of TGF-β1-mediated suppression was dependent on the dose of bFGF used. In addition, we noted that the inclusion of neutralizing antibody to TGF-β1 significantly augmented the clonogenic response to GM-CSF. We have also shown that 10 ng/mL or 100 ng/mL of bFGF resulted in a 30% to 100% increase in GM-CSF-mediated progenitor cell growth (P= .0001). These data suggest that bFGF may enhance myelopoiesis by modulating the inhibitory response to TGF-β1.

The prothoracicotropic hormone (PTTH), which stimulates ecdysteroid synthesis in the prothoracic glands, is produced, in the dorso-lateral protocerebrum of Manduca sexta, by paired peptidergic neurons, the lateral neurosecretory cell group III (L-NSC III). Our study revealed ultrastructural features of L-NSC III, identified by immunogold labeling, and compared developing and diapause states. In developing and early-diapause pupae, L-NSC III soma ultrastructure is similar and is characterized by numerous clusters of neurosecretory granules (NSG) and an extensive trophospongium formed by satellite-glial cells. However, as diapause progresses, the ultrastructure changes, with the NSG becoming concentrated into large clusters separated by highly organized rough endoplasmic reticulum. Most conspicuous is a substantial reduction in the number of Golgi complexes and the glial trophospongium, and the presence of stacked plasma membrane separating the glia and neuron somata. The deep-diapause soma also has abundant glycogen deposits and autophagic vacuoles. With diapause termination, this morphology reverts to the nondiapause ultrastructure within three days, i.e. just before PTTH release that evokes development to the adult. During PTTH release the abundance of NSG in the soma does not change, suggesting that NSG depletion in the perikarya is not a marker for neurosecretion by the L-NSC III.

The prothoracicotropic hormones (PTTHs) are cerebral peptides that control insect postembryonic development by stimulating the prothoracic glands to synthesize ecdysteroids. InManduca sexta, the tobacco hornworm, two classes of PTTH are distinguished by their Mr, small (ca. 7 kDa) and big PTTH (ca. 25–30 kDa). Little is known about the physical nature of the PTTHs and this study takes a first step towards defining characteristics of theManducabig PTTH. The neurohormone has a Stokes radius of 2.59 nm and a sedimentation coefficient of 2.76 S. Based on these data, an Mrof 29,443.7 and anf/f0of 1.27 were calculated. Combined, the physical data revealManducabig PTTH is an asymmetrical acidic homodimeric peptide with intra- and intermolecular disulfide bonds.

Approximately two-thirds of the total amount of ecdysteroids in late—pharate adults of the wax moth, Galleria mellonella, were found in the ovaries and one-third in the ovariectomized body. Chemical analysis of these ecdysteroids by thin-layer and high-pressure liquid chromatography, coupled with an ecdysteroid radioimmunoassay, revealed the presence of 2-deoxyecdysone, ecdysone and 20-hydroxyecdysone, as well as high and low polarity unknowns. The predominant identifiable ecdysteroid in both the ovaries and ovariectomized body was 2-deoxyecdysone, followed by lesser amounts of ecdysone and 20-hydroxyecdysone, respectively. Incubation of late-pharate adult ovaries in culture medium revealed that they synthesize and secrete ecdysteroids in vitro. The in vitrodistribution of ecdysteroids between ovaries and incubation medium was similar to that observed between ovaries and ovariectomized bodies in situand the predominant identifiable moiety both retained and released by the ovaries in vitrowas 2-deoxyecdysone, followed by lesser amounts of ecdysone and 20-hydroxyecdysone. Collectively, these results support the idea that the ecdysteroids synthesized by the ovaries of late-pharate adult Galleriaare both stored and secreted and that the quantity of a specifically secreted ecdysteroid is precisely controlled. This apparent regulation of the distribution of ovarian ecdysteroids raises the possibility that the stored and secreted forms have distinct functions in the reproductive physiology of this insect.

An in vitroassay for the prothoracicotropic hormone has been developed that utilizes an ecdysone radioimmunoassay to quantify the increase in the rate of ecdysone synthesis elicited by the neurohormonal activation of the prothoracic glands. The rapidity, reproducibility, and accuracy of the assay were maximized by using one member of a gland pair as the control and the other as the test gland. This was possible because the basal rates of ecdysone synthesis by the members of a gland pair were equivalent. Activation was demonstrated to be dose dependent and specific, with prothoracicotropic hormone activity present only in homogenates of brain. The in vitroactivation of the prothoracic glands was verified with the Manducabioassay for the prothoracicotropic hormone in which the morphological responses to the hormone were correlated with increased in vivoecdysone titers. These results provide unequivocal evidence that the activation of the prothoracic glands by the prothoracicotropic hormone is direct and suggest that activation represents an increase in a basal rate of ecdysone synthesis.

Pupal development is elicited early in the last larval instar of the tobacco hornworm, Manduca sexta (Johannson), by a precise temporal and quantitative increase in the haemolymph titre of 20-hydroxyecdysone. This increase in the titre is referred to as the pupal commitment peak, and it occurs once the titre of juvenile hormone (JH) has dropped. If the haemolymph titre of JH remains elevated at this time due to topical application of the hormone or of its analogue ZR512, commitment is delayed or inhibited in a dose-dependent manner. This delay or inhibition is due to the curtailment of the commitment peak in the ecdysteroid titre, which results from a failure of the prothoracic glands (PG) to increase the synthesis/secretion of the premoulting hormone, ecdysone. Since the PG from ZR512- and JH 1-treated larvae are capable of being activated in vitro by the prothoracicotropic hormone (PTTH), the effect of JH on the PG does not involve suppression of gland sensitivity to PTTH. The locus of the JH effect was determined to be the brain-retrocerebral complexes (Br-CC-CA), on the basis of experiments which tested the effect of implanted Br-CC-CA from pre-commitment larvae treated with JH on the occurrence of pupal commitment in head-ligated larval hosts. The implanted, JH-treated Br-CC-CA exhibited a delayed release of PTTH, and the effect was at concentrations of JH that were physiological. These results argue that JH functions to control the time during the last larval instar when pupal commitment occurs by dictating when PTTH will undergo gated release.

Prothoracic glands from diapausing pupae of the tobacco hornworm, Manduca sexta, synthesize markedly less ecdysone in vitro in response to prothoracicotropic hormone (PTTH) than do glands from non-diapausing pupae. Impaired steroidogenesis is also observed in glands from diapausing animals exposed to agents that enhance ecdysone synthesis in non-diapausing pupal glands by increasing intracellular levels of cAMP (l-methyl-3-isobutylxanthine, dibutyryl cAMP, and the calcium ionophore A23187). In contrast, prothoracic glands from diapausing pupae synthesize significantly more cAMP in response to PTTH and A23187 than do those from non-diapausing pupae. These observations indicate that the PTTH-refractoriness characteristic of prothoracic glands during diapause results from a lesion in the steroidogenic pathway occurring beyond the level of the PTTH receptor-adenylate cyclase system. The diapause condition of the prothoracic glands (reduced ecdysone synthesis accompanied by enhanced cAMP formation) can be mimicked by extirpation of the brain of a non-diapausing pupa. Thus, cellular changes in the prothoracic glands associated with diapause may arise as a result of the absence of some factor produced by the pupal brain (e.g. PTTH).

Pupal diapause in the tobacco hornworm, Manduca sexta (Johannson), is characterized by the absence of an increased ecdysteroid titre in the haemolymph during the first week of pupal life as measured by radioimmunoassay. This virtual absence of the steroid moulting hormone is thought to be responsible for the diapause state and it is apparently a consequence of the failure of the diapausing pupal prothoracic glands (PG) to synthesize ecdysone at an increased rate. In Manduca, this apparent failure of PG activation during diapause may be in response to two endocrinological circumstances: the curtailment of PTTH release as opposed to its synthesis and transport, and the development of refractoriness of the gland to stimulation by PTTH. The former was examined by measuring PTTH activity present in brains and brain-retrocerebral complexes of diapausing and non-diapausing pupae and the latter by assessing PG biosynthetic activity in vitro in the presence and absence of PTTH.

A larval moult in the tobacco hornworm, Manduca sexta, involves an endocrine cascade that begins with the release of a cerebral peptide hormone, the pro-thoracicotropic hormone (PTTH). The release of PTTH is gated, occurs during the scotophase and appears to be developmentally cued. In fourth instar Manduca larvae, PTTH release into the haemolymph occurs as a single burst over a few hours during the head critical period, i.e. the time during which the head (brain) is needed for the initiation of the moult to the fifth (last) instar. Released PTTH activates the prothoracic glands (PGs), and within a few hours the cumulative effect of this event results in a dramatic increase in the haemolymph ecdysteroid titre, which then elicits the moult. An assessment of the capacity of the corpora allata (CA) to synthesize juvenile hormone (JH) in vitro indicates that the above sequence of endocrine events begins only when JH synthesis has reached a nadir for the instar. Since CA activity is an indirect measure of the haemolymph titre of the hormone, it is conceivable that the developmentally cued release of PTTH is permissively controlled by a decreasing haemolymph titre of JH. With the increase in the ecdysteroid titre which marks the end of this endocrine cascade, the CA again become active, presumably to cause the increase in the JH haemolymph titre which directs the larval moult. This in-vestigation has thus established the temporal and quantitative dynamics of the PTTH-PG axis that drive larval moulting and provides insight into the inter-endocrine regulatory relationships that may exist between the ecdysteroids and JHs. These possible relationships and the role of the brain in their regulation are discussed.

A recently isolated haemolymph protein appears to be an important regulator of ecdysone biosynthesis by prothoracic glands in Manduca sexta. Using a dose-response titration protocol, the haemolymph titre of this stimulatory factor was determined during the last larval instar. The titre was high (>2·0UmU1) on days0 and 1, then dropped significantly to 0·55 U ml-1 on day 2, and remained depressed until day 4. The titre of the stimulatory factor then increased to a peak of 1·62U ml-1 on day 7, and remained elevated (approx. 1·1 U ml-1) until the end of the instar. A set of physical and biochemical criteria was used to confirm that the stimulatory activity present in haemolymph on different days of the instar rep-resented the presence of the factor. The data are consistent with the hypothesis that fluctuations in the titre of the haemolymph stimulatory factor play a critical role in regulating ecdysone biosynthesis during larval-pupal development.

Pupal diapause in the tobacco hornworm Manduca sexta is a photoperiodically induced event expressed by the absence of release of the cerebral peptide prothoracicotropic hormone. A short-day photoperiod (light-dark, 12:12) during larval development programs the brain to forego the release of prothoracicotropic hormone in the pupal stage. Prothoracicotropic hormone release can occur in the pupae and diapause can be averted if developing larvae experience an intervening long-day photoperiod (light-dark, 16:8) for 3 days early in the last larval instar. Implanted brains from larvae that received such long-day stimulation can reverse the diapause commitment of short-day-reared diapause-destined larvae, thus overriding the diapause program of the recipient. A system was established that demonstrated that this photoperiodic reprogramming of the insect's brain occurs in vitro. Brain-retrocerebral complexes from day 1 instar larvae were cultured in vitro for 3 days under long-day conditions. The effect of this non-diapause photoperiod on the diapause program of these brain-retrocerebral complexes was assessed by assaying the effect of the brains from these cultures on the incidence of non-diapause development in diapause-destined larval hosts. Brains that received a long-day photoperiod reversed the pupal diapause program of the recipient larvae, whereas brains that received a short-day photoperiod were ineffective. Therefore, the larval brain-retrocerebral complex of this insect is capable of photoperiodic light reception in vitro, in that it can sense the number of light-dark cycles to which it is exposed, store this environmental input, and express the information 9 days later at another stage of development. These findings demonstrate that the photoperiodic clock of an animal can be reprogrammed by a light-dark cycle in vitro.

The chemical basis of an ant-seed interaction was investigated for the antAphaenogaster rudis and the ant-dispersed violetViola odorata. A laboratory behavioral bioassay was developed to chemically identify the attractant responsible for the interaction. The ant attractant, localized in the elaiosome, was classified as a lipid by both field and laboratory bioassays. Assays of partially purified lipid extracts revealed that the principal attractant may be a diglyceride. Gas-liquid chromatography analysis of the hydrolyzed diglyceride fraction revealed oleic acid as the major fatty acid present, suggesting that either 1,2- or 1,3-diolein may be the attractant. Structure-activity correlations for lipid standards demonstrated a clear preference for the diglyceride 1,2-diolein. The data also suggest that ricinoleic acid is not the lipid eliciting the ant response toViola odorata, as had been previously suggested.

The kinetics of prothoracicotropic hormone (PTTH) activation of pupal prothoracic glands (PG) of the cabbage army worm,Mamestra brassicae, and silkworm,Bombyx mori, were determined in vitro. Activation was assessed by comparing the increase in the rate of ecdysone synthesis by one member of a PG pair incubated with a PTTH preparation from pupal brains with the basal rate of synthesis of the other PG incubated without PTTH. A time course of ecdysone synthesis revealed thatBombyxPTTH extract activatedBombyxPG, andMamestraPTTH extract activatedMamestraPG. Dose responses of activation ofBombyxandMamestraPG by their respective PTTH were saturable and were indicative of neurohormonal activation. TheBombyxPG were half-maximally activated (A50) by far less PTTH thanMamestraPG, 0.34 and 0.91 brain equivalents, respectively. Heterologous dose response of activation studies, in which PTTH and PG fromMamestra, Bombyxand the tobacco hornworm,Manduca sexta, were assayed for interspecific PG sensitivity and PTTH specificity, revealed cross-reactivity among the three PTTH-PG axes, withManducaPG being more sensitive to the PTTH of the other species andBombyxPTTH being the most effective in activating the PG of the other two species.

The distribution of prothoracicotropic hormone in the pupal brain of Manduca sextahas been determined by an in vitroassay for prothoracic gland activation. Prothoracicotropic activity was observed in both the brain and retrocerebral complex, but predominantly in the dorosolateral regions of the protocerebrum. Of the two groups of neurosecretory cells present in this area of the brain, only the two lateral type III neurosecretory cells exhibited significant prothoracicotropic hormone activity. Further analysis revealed that the neurohormone was localized in only one of the two type III cells, suggesting that a single neurosecretory cell in each hemisphere is the source of the hormone at the stage examined (day 0). Prothoracicotropic hormone activity was detected in both the corpora allata and the corpora cardiaca, but the corpora allata contained 2 to 9 times the activity of the corpora cardiaca, depending on developmental stage. The significantly higher level of activity in the corpora allata suggests that they may be the neurohemal organs through which the prothoracicotropic hormone of Manducais released.

Prothoracic glands of tobacco hornworm larvae cultured in vitrosecrete into the culture medium a substance which was active in ecdysone bioassays and determined to be ecdysone-like by radioimmunoassay. The prothoracic glands appear to be the sole source of this substance. The material was identified as α-ecdysone by thin-layer, gas-liquid, and high-resolution liquid chromatography and mass spectrometry. No other active moulting principles were present in the medium. It appears that the prothoracic gland secretes a prohormone, α-ecdysone, which is subsequently converted into the active moulting hormone, β-ecdysone, in other insect tissues.

Discusses ways for optimizing document-intensive processes (DIP) in financial institutions. Production processes; Archival processes; Classification by origin; Understanding document technology.

Focuses on the incorporation of employee enhancement into a bank's culture. Factors that contribute to employee's low morale; Importance of communication in enhancing employee performance; Participation of employees in bank management.