Two unusual complications of neuroleptic malignant syndrome.

Neuroleptic malignant syndrome (NMS) is a rare but well described complication of the administration of antipsychotic agents. Compartment syndrome, with increased pressures within the confined space of fascial sheaths leading to compression damage of the contained tissue, similarly is well described. Brachial plexus injuries caused by patient malposition are also very rare but a few cases have been reported. We report a case where these three complications occurred together. This was attributable to the patient developing NMS whilst asleep in the prone position overnight.

Application of specific ELISAs for BMP15 and GDF9 to cumulus cell extracts from infertile women.

Bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9) are oocyte-specific paracrine factors which regulate ovarian cumulus cell (CC) functions. This study aimed to investigate if BMP15 and GDF9 bound to CCs can be characterized, quantified, and show an association with IVF outcomes in infertile women. BMP15 and GDF9 ELISAs were validated and applied to discarded CC extracts. Pooled CCs from individual patients were collected from 120 (cohort 1; BMP15 only) and 81 infertility patients (cohort 2; BMP15 and GDF9) undergoing superovulation. BMP15 and GDF9 levels expressed per CC DNA were correlated with maternal age, clinical and embryology data. Total BMP15 and GDF9 were highly correlated with each other (r = 0.9, p < 0.001). The GDF9:BMP15 ratio was unrelated to oocyte number or age. BMP15/CC DNA and GDF9/CC DNA were unaffected by the type of superovulation and were not related to oocyte/embryo outcomes.

The impact of breast cancer surgery on functional status in older women - A systematic review of the literature.

Primary endocrine therapy as treatment of breast cancer is only recommended in older women with limited life expectancy. However, many older women opt for endocrine therapy due to concerns regarding frailty and potential decline in function after surgery. A decline in functional status after surgery is documented in some cancer types, such as colorectal, however, the full impact of breast cancer surgery is less understood. A systematic review was performed to examine the evidence for impact of breast cancer surgery on functional status in older women. PubMed and Embase databases were searched. Studies were eligible if performed within the last 10 years; included patients over the age of 65 years undergoing breast cancer surgery; included stratification of results by age; measured functional status pre-operatively and at least six months following surgery. A total of 11 studies including 12 030 women were appraised. Two studies represented level-II and nine level-IV evidence. Overall, physical activity level was negatively impacted by breast cancer surgery and this was compounded by the extent of surgery. Evidence for impact of breast cancer surgery on quality of life, fatigue and cognition, was conflicting. The possibility of decline in functional status after breast cancer surgery should be discussed in all older women considering surgery. A structured exercise program may improve the negative effects of surgery on physical activity. Further work is required in the areas of quality of life, fatigability and cognition.

Non-canonical cyclic AMP SMAD1/5/8 signalling in human granulosa cells.

Development of mammalian ovarian follicles is promoted by the combined action of endocrine cues and paracrine factors. Follicle stimulating hormone (FSH), through the action of cAMP drives follicular growth and development. The oocyte secretes powerful growth factors such as bone morphogenetic protein 15 (BMP15) to regulate granulosa cell proliferation, metabolism, steroidogenesis and differentiation through the activation of SMAD1/5/8. This study investigated the role of the cAMP signalling pathway on SMAD1/5/8 action in human granulosa cells. Cyclic AMP enhanced BMP15-induction of a SMAD1/5/8-specific BRE reporter. Moreover, in the absence of BMP ligand, cAMP also activated SMAD1/5/8-induced BRE activity. Cyclic AMP increased canonical downstream targets of BMP signalling such as inhibitor of differentiation (ID) mRNA expression. The observed effects were not mediated by secretion of BMPs as cAMP did not promote BMP ligand mRNA expression and a BMP extracellular antagonist, the BMP type II receptor ectodomain, did not affect cAMP-induced ID mRNA expression. Finally, the ERK1/2 pathway was shown to be required for the maintenance of cAMP-induced SMAD1/5/8 activity. Together our results suggest a novel and non-canonical pathway for cAMP signalling in human granulosa cells. Cyclic AMP appears to promote SMAD1/5/8 pathway activity intracellularly and has the ability to activate canonical SMAD1/5/8 downstream targets. Our results add another layer of complexity to the interactions between endocrine signalling and oocyte-secreted BMP ligands during folliculogenesis. Given the importance of both cAMP and SMAD1/5/8 pathways in follicular development, these interactions are likely required for the fine-tuning of oocyte paracrine signalling by endocrine stimuli.

Role of calcineurin in the regulation of human lung mast cell and basophil function by cyclosporine and FK506.

BACKGROUND AND PURPOSE: Cyclosporine and FK506 are thought to act by targeting the Ca2+-dependent protein phosphatase, calcineurin. The aim of the present study was to determine whether cyclosporine and FK506 stabilize mast cells and basophils by interacting with calcineurin. EXPERIMENTAL APPROACH: The effects of cyclosporine and FK506 on the IgE-mediated release of histamine from mast cells and basophils were evaluated. The presence of calcineurin in cells was determined by Western blotting. Ca2+-dependent protein phosphatase activities were assessed in cell extracts using a synthetic phosphorylated peptide that is known to serve as a substrate for calcineurin. KEY RESULTS: FK506 was about 100-fold more potent than cyclosporine as an inhibitor of IgE-dependent histamine release from mast cells and basophils. Immunoblotting of solubilized preparations of purified cells demonstrated the presence of calcineurin in mast cells and basophils. In enzyme assays, mast cells expressed approximately 7-fold higher Ca2+-dependent protein phosphatase activity than basophils. Whereas cyclosporine effectively inhibited Ca2+-dependent protein phosphatase activity in cell extracts, FK506 was considerably less effective. CONCLUSIONS AND IMPLICATIONS: FK506 and cyclosporine inhibit the stimulated release of histamine from mast cells and basophils. However, the ability of cyclosporine, but not FK506, to inhibit Ca2+-dependent protein phosphatase activity questions whether FK506 stabilizes mast cells and basophils by interacting with calcineurin.

The myosin light chain enhancer and the skeletal actin promoter share a binding site for factors involved in muscle-specific gene expression.

The myosin light chain (MLC) 1/3 enhancer (MLC enhancer), identified at the 3' end of the skeletal MLC1/3 locus, contains a sequence motif that is homologous to a protein-binding site of the skeletal muscle alpha-actin promoter. Gel shift, competition, and footprint assays demonstrated that a CArG motif in the MLC enhancer binds the proteins MAPF1 and MAPF2, previously identified as factors interacting with the muscle regulatory element of the skeletal alpha-actin promoter. Transient transfection assays with constructs containing the chloramphenicol acetyltransferase reporter gene demonstrated that a 115-bp subfragment of the MLC enhancer is able to exert promoter activity when provided with a silent nonmuscle TATA box. A point mutation at the MAPF1/2-binding site interferes with factor binding and abolishes the promoter activity of the 115-bp fragment. The observation that an oligonucleotide encompassing the MAPF1/2 site of the MLC enhancer alone cannot serve as a promoter element suggests that additional factor-binding sites are necessary for this function. The finding that MAPF1 and MAPF2 recognize similar sequence motifs in two muscle genes, simultaneously activated during muscle differentiation, implies that these factors may have a role in coordinating the activation of contractile protein gene expression during myogenesis.

Identification of specific inhibin A-binding proteins on mouse Leydig (TM3) and sertoli (TM4) cell lines.

The binding of human inhibin A to cell surface binding proteins of mouse Leydig (TM3) and Sertoli (TM4) cell lines was investigated. Scatchard analysis identified two classes of inhibin A-binding sites on TM3 (K(d(1)) = 85 pM and 4,160 sites/cell; K(d(2)) = 520 pM and 12,500 sites/cell) and TM4 (K(d(1)) = 61 pM and 2,620 sites/cell; K(d(2)) = 520 pM and 10,400 sites/cell) cells. Compared with inhibin A, inhibin B only partially competed [(125)I]inhibin A binding (6-8%), whereas activin A competed weakly (<0.01%). Chemical cross-linking of [(125)I]inhibin A to both cell lines identified five [(125)I]inhibin A binding complexes with apparent molecular masses of 70, 95, 145, 155, and more than 200 kDa. Inhibin A displacement of [(125)I]inhibin A from each of these cross-linked species (ED(50) = 60-110 pM) closely resembled displacement from intact TM3 (ED(50) = 97 +/- 32 pM) and TM4 (ED(50) = 75 +/- 28 pM) cells, suggesting that all of these proteins are involved in the high affinity inhibin A binding complex. Immunoprecipitation of iodinated inhibin A complexed to TM3 and TM4 cells with an antibody against human betaglycan identified protein complexes of more than 200, 145, and 95 kDa. It is concluded that the high affinity binding complex for inhibin A found in these cell lines consists of betaglycan and several proteins of unknown identity and may represent the putative inhibin receptor complex.

Inhibin binding sites and proteins in pituitary, gonadal, adrenal and bone cells.

Activin signals via complexes of type I (50-55 kDa) and II (70-75 kDa) activin receptors, but the mechanism of inhibin action is unclear. Proposed models range from an anti-activin action at the type II activin receptor to independent actions involving putative inhibin receptors. Two membrane-embedded proteoglycans, betaglycan and p120, have recently been implicated in inhibin binding, but neither appears to be a signalling receptor. The present studies on primary cultures of rat pituitary and adrenal cells, and several murine and human cell lines were undertaken to characterise inhibin binding to its physiological targets. High affinity binding of inhibin to the primary cultures and several of the cell lines, like that previously described for ovine pituitary cells, was saturable and reversible. Scatchard analysis revealed two classes of binding sites (K(d) of 40-400 and 500-5000 pM, respectively). Affinity labelling identified [125I]inhibin binding proteins with apparent molecular weights of 41, 74, 114 and >170 kDa in all cell types that displayed high affinity, high capacity binding of inhibin. Additional labelling of a 124 kDa species was evident in gonadal TM3 and TM4 cell lines. In several cases, activin (> or =20 nM) competed poorly or not at all for binding to these proteins. The 74, 114 and >170 kDa inhibin binding proteins in TM3 and TM4 cells were immunoprecipitated by an anti-betaglycan antiserum. These three proteins correspond in size to the activin receptor type II and the core protein and glycosylated forms of betaglycan, respectively, that have been proposed to mediate anti-activin actions of inhibin, but the identity of the 74 kDa species is yet to be confirmed. Studies of [125I]inhibin binding kinetics and competition for affinity labelling of individual binding proteins in several cell lines suggest these three species and the 41 and 124 kDa proteins form a high affinity inhibin binding complex. In summary, common patterns of inhibin binding and affinity labelling were observed in inhibin target cells. Novel inhibin binding proteins of around 41 and 124 kDa were implicated in the high affinity binding of inhibin to cells from several sources.

Recombinant and cellular expression of the murine chemotactic protein, CP-10.

The S100 protein CP-10 (chemotactic protein, 10 kD), a potent chemotactic factor for murine and human polymorphonuclear cells (PMN) and murine monocytes, has been purified in small amounts from supernatants of activated murine spleen cells (Lackmann et al., 1992). To obtain a more abundant source of the protein, CP-10 was expressed in Escherichia coli as a fusion protein with glutathione S-transferase (GST). The property of S100 proteins to undergo calcium-dependent conformational changes was used in a novel approach to optimize the release of recombinant (r) CP-10 by thrombin cleavage. Purified rCP-10 was characterized by amino-terminal sequence analysis and bioassays. Optimal chemotactic activity of rCP-10 for murine PMN and WEHI-265 monocytoid cells was 10(-11) M (native protein has optimal chemotactic activity between 10(-11) and 10(-13) M). Immunization of rabbits with the GST/CP-10 fusion protein bound to glutathione-agarose beads resulted in high titer, specific antibodies that neutralized CP-10-initiated chemotaxis and were suitable for immunoblotting. A combination of Western and Northern analyses identified CP-10 in murine peritoneal exudate PMN and macrophages, splenocytes, bone marrow cells, and WEHI-265 cells (all of myeloid origin), but not in thymus, liver, lung, 3T3 fibroblasts, EL4 lymphoma cells, or bEND 3 brain endothelial cells, indicating cell-specific regulation of CP-10 expression.

Effect of rioprostil on aspirin-induced gastrointestinal mucosal changes in normal volunteers.

Rioprostil, a 15-deoxy-16-methyl prostaglandin E1, was evaluated for its effect on aspirin-induced gastrointestinal mucosal changes in normal volunteers. Fifty-six normal male volunteers were evaluated by endoscopy in a double-blind, placebo-controlled study. Aspirin was given at a dose of 975 mg four times per day. Rioprostil was given at doses of 60, 120, and 300 micrograms four times per day. Rioprostil, at both antisecretory and subantisecretory doses, prevented or reduced aspirin-induced injury. Increased stool frequency was the most common side effect and appeared to be a dose-related effect of rioprostil occurring at only antisecretory doses.

Reduced locomotor activity as an acute effect of damage to superior colliculus in rats.

Rats with either electrolytic or radiofrequency lesions of the superior colliculus were tested in an open-field within 24 h of operation. They crossed significantly fewer squares and spent more time motionless then control animals, an effect that disappeared upon retesting 13 days later. Previously reported locomotor hyperactivity thus appears to be a chronic but not an acute effect of collicular damage in rats.

Immunodetection of the murine chemotactic protein CP-10 in bleomycin-induced pulmonary injury.

The murine S-100 protein designated CP-10 is a potent chemotactic factor for phagocytic cells, exhibiting optimal activity in the picomolar range. We assessed the role of this cytokine in the inflammatory response to pulmonary injury following intratracheal administration of bleomycin to mice. In the lungs of normal animals, strong cytoplasmic immunostaining for CP-10 was demonstrable in all recognisable neutrophil leucocytes sequestered within alveolar capillaries. Following induction of pulmonary inflammation in susceptible C57BL/6 mice, numerous CP-10-positive neutrophils were observed, but many of the recruited neutrophils did not exhibit staining for CP-10. No other cells were immunoreactive. The concentration of CP-10 in bronchoalveolar lavage (BAL) fluids from normal mice and mice administered intratracheal saline was below the level of detection by enzyme immunoassay. In contrast, nanomolar levels of CP-10 were detected in unconcentrated BAL fluids from C57BL/6 mice after bleomycin-induced injury, and the presence of monomeric CP-10 was demonstrable by Western blotting. Elevation of CP-10 levels correlated with the influx of inflammatory cells in C57BL/6 mice, but was not demonstrable in BAL fluids from BALB/c mice, which are resistant to pulmonary injury by bleomycin. We conclude that CP-10 may contribute to the recruitment of inflammatory cells in bleomycin-induced lung damage.

Family caregivers' criticism of patients with schizophrenia.

OBJECTIVE: Relationships were examined between patients' negative symptoms, family caregivers' knowledge of schizophrenia, caregivers' attributions about the cause of patients' symptoms, and caregivers' response to the symptoms. METHODS: A sample of 84 caregivers of patients with schizophrenia in Brisbane, Australia, were interviewed using a structured format and measures designed for the study. RESULTS: Results of regression analyses indicated that three variables significantly predicted caregivers' criticism of patients--a smaller proportion of negative symptoms in the patient's overall symptom pattern, the caregiver's low level of knowledge about the illness, and the caregiver's attributing the cause of negative symptoms to the patient's personality rather than to the illness. CONCLUSIONS: Overall, findings supported the utility of an attributional framework in enhancing conceptions about and research on schizophrenia and family caregiving.

The use of Hickman-Broviac catheters for paediatric radiotherapy.

We describe the first use of Hickman-Broviac catheters in 20 children under the age of 4 years for repeated daily general anaesthesia for radiotherapy. These children received a total of four hundred and eighty two general anaesthetics. The advantages and disadvantages of using Hickman-Broviac lines are discussed.

The effects of cosmic particle radiation on pocket mice aboard Apollo XVII: VI. launch, flight, and recovery.

The final phase to fly five pocket mice in the Apollo XVII command module was carried out at the NASA Kennedy Space Center. Upon completion of the 13-d space flight, the package was removed from the spacecraft and, after having been purged with an oxygen-helium gas mixture, was flown to American Samo. Four of the five mice were recovered alive from the package. Analysis of the mouse that died during the flight revealed several factors that could have contributed to its death, the chief of which was massive hemorrhage in its middle ear cavities.

The U.S. study of work incapacity and reintegration.

In many countries, including the United States, the number of persons being awarded long-term or permanent disability benefits has risen dramatically in recent years. Government agencies, advocates for the disabled, and others are looking for ways to help persons with disabilities return to the labor force. The Work Incapacity and Reintegration (WIR) Study was developed to address that issue. The United States and five other countries--Germany, Denmark, Sweden, Israel, and the Netherlands--have participated in a cross-national study of work incapacity under the auspices of the International Social Security Association. The study had two objectives: to examine the factors that influence the pattern of work resumption among persons disabled by a back condition and to identify the medical and nonmedical interventions that are most effective in helping such persons reenter the labor force. Samples for the U.S. national study were drawn from four cohorts: Social Security Disability Insurance (DI) beneficiaries, Supplemental Security Income (SSI) beneficiaries, and recipients of temporary disability insurance (TDI) benefits from the states of California and New Jersey. Only the TDI recipients were included in the comparative study. This article discusses the study design and methodology and summarizes the findings of the U.S. national study. Findings from the U.S. study show significant differences between the two cohorts in terms of work resumption and other characteristics. The proportions of respondents from the TDI cohorts who were working at the third and final study contact ranged from 53 percent to 65 percent, compared with less than 5 percent of the DI and SSI respondents. Respondents from the DI and SSI cohorts were on average about 10 years older than the TDI respondents, were less well educated, and reported more physical demands in their usual work. They also reported lower levels of functional capacity, higher levels of pain, and a much greater tendency to have other chronic illnesses. The types of medical treatments provided were remarkably uniform across cohorts and, within cohorts, between those who did and did not resume working. Thus, no medical intervention was identified that showed a significantly higher success rate in terms of facilitating a return to work. However, changes made in the work environment by the employer were an important factor in work reintegration; about 80 percent of respondents who resumed working did so with the help of workplace accommodations. In addition, since respondents with fewer physical demands in their job were more likely to return to work, there appears to be some potential for job retraining as a means of promoting a return to work. The Social Security Administration should consider these findings in developing strategies to help disabled workers reenter the labor force.

Inhibin and premature ovarian failure.

BACKGROUND: Elucidation of the causes of premature ovarian failure (POF) is difficult due to the heterogeneity of the condition. Inhibin is a potential candidate gene for POF based on its dual actions on FSH secretion by the pituitary and gametogenesis in the gonads. A missense mutation in the inhibin alpha subunit gene (INHA G769A) is associated with POF in several populations. However, there is phenotypic heterogeneity in INHA G769A mutation carriers. METHODS: Relevant studies were identified by searching PubMed and mutational frequencies combined for meta-analysis. RESULTS: Meta-analysis of published studies revealed a risk difference of 0.04 (-0.030 to 0.11). The occurrence of asymptomatic carriers in populations suggests incomplete penetrance and/or a multi-genetic cause of POF. We propose that a decline in inhibin bioactivity caused by the mutation could increase FSH levels; and in a susceptible individual, the heightened sensitivity to gonadotrophins causes POF. Impaired paracrine effects of inhibin could impact folliculogenesis due to reduced antagonism of activin, bone morphogenetic protein 15 and growth differentiation factor 9. Functional studies of this mutation indicate normal production of dimeric inhibin A and B and impaired bioactivity of inhibin B. CONCLUSIONS: The identification of an autosomal mutation in the inhibin alpha subunit gene that is significantly linked to POF in certain ethnic populations highlights the role of inhibin in the regulation of ovarian biology and fertility. Although the reduction of inhibin B bioactivity by the INHA G769A mutation is clearly not the only cause, evidence suggests that this change may serve as a susceptibility factor, increasing the likelihood of POF.

Transglutaminase inhibitors induce hyperproliferation and parakeratosis in tissue-engineered skin.

BACKGROUND: The transglutaminase (TG) family consists of eight distinct isoforms. TG types 1, 3 and 5 play a major role in normal skin development, with TG2 also being elevated during dermal wounding. TG1, 3 and 5 are responsible for the cross-linking of keratin precursors and formation of the cornified envelope during keratinocyte differentiation. TG2 may play a role in keratinocyte basement membrane cross-linking. Abnormal TG expression has been demonstrated in Darier disease, Netherton syndrome, psoriasis and lamellar ichthyosis. During a recent investigation of skin contraction in tissue-engineered skin, transglutaminase inhibitors were found to produce hyperproliferation and parakeratosis. OBJECTIVES: Accordingly, this study was designed to study the effect of pan-transglutaminase inhibition on morphology of tissue-engineered skin and expression of keratinocyte differentiation and proliferation-associated antigens. METHODS: We used a tissue-engineered model of human skin, based on de-epidermized acellular human dermis, seeded with normal keratinocytes and dermal fibroblasts and cultured at an air-liquid interface. The pan-transglutaminase inhibitors putrescine, NTU283 (1-dimethyl,2-[(oxopropyl)thio]imidazolium) and NTU285 (N-benzyloxycarbonyl-l-glutaminyl-6-dimethylsulfonium-5-oxo-l-norleucine) were added to the culture medium. After 28 days, histology and immunohistochemistry for collagen IV, involucrin and cytokeratins 6, 10 and 16 were performed. RESULTS: Keratinocyte hyperproliferation and parakeratosis were seen in response to transglutaminase inhibition. Inhibition of transglutaminase also resulted in loss of basement membrane collagen IV. Involucrin and cytokeratins 6 and 16 were confined to the basal layers in control composites but expressed throughout the epidermis in response to transglutaminase inhibition. A distinct band of expression of cytokeratin 10 was seen in the upper stratum granulosum of control composites but only patchy expression was seen after transglutaminase expression. CONCLUSIONS: Pan-transglutaminase inhibition inhibits terminal differentiation of keratinocytes, leading to a hyperproliferative epidermis with parakeratosis and enhanced expression of involucrin and cytokeratins 6 and 16. Expression of the differentiation-associated cytokeratin, cytokeratin 10, is reduced. Basement membrane integrity is also lost as a result of transglutaminase inhibition.

Investigation of keratinocyte regulation of collagen I synthesis by dermal fibroblasts in a simple in vitro model.

BACKGROUND: Hypertrophic scarring and skin graft contracture are major causes of morbidity after burn injuries. A prominent feature is excessive fibroplasia with accumulation of increased fibrillar collagen relative to normal scar tissue. The application of split-thickness skin grafts or cultured epithelial autografts to burn wounds is known to reduce scarring and contraction. OBJECTIVES: To investigate further how the keratinocyte influences underlying fibroblast behaviour by examining the influence of keratinocytes on fibroblast collagen synthesis, using a new assay for collagen synthesis never previously applied to skin cell biology. METHODS: We investigated the influence of the keratinocyte on fibroblast synthesis of type I collagen using an immunoassay for the aminoterminal propeptide of type I collagen (P1NP) in conditioned medium from monocultures and cocultures of keratinocytes and fibroblasts over 14 days. The importance of the physical presence of the keratinocyte was investigated by comparing cocultures of keratinocytes and fibroblasts against fibroblast monocultures with keratinocyte-conditioned medium. Pharmacological agents known to promote fibroblast proliferation [basic fibroblast growth factor (bFGF)], keratinocyte proliferation [insulin-like growth factor (IGF)-1], modify scarring in vivo[tumour necrosis factor (TNF)-alpha] or modify collagen biochemistry [putrescine, estrone, estradiol and beta-aminopropionitrile (beta-APN)] were then investigated for their effect on collagen synthesis in fibroblasts and in keratinocyte/fibroblast cocultures. RESULTS: Keratinocytes in coculture with fibroblasts, and keratinocyte-conditioned medium, both reduced fibroblast P1NP synthesis. Of the pharmacological agents investigated, bFGF, IGF-1, TNF-alpha and beta-APN all increased collagen synthesis both in monocultures of fibroblasts and in cocultures of keratinocytes and fibroblasts. CONCLUSIONS: Fibroblast collagen synthesis appears to be downregulated by keratinocyte-derived cytokines. Fibroblast growth factors and proinflammatory cytokines appear to be able partially to overcome this downregulation and to increase collagen synthesis.