Decision making on timing of surgery for hip fracture patients on clopidogrel.

Patients taking clopidogrel who sustain a fractured neck of femur pose a challenge to orthopaedic surgeons. The aim of this study was to determine whether delay to theatre for these patients affects drop in haemoglobin levels, need for blood transfusion, length of hospital stay and 30-day mortality. A retrospective review of all neck of femur patients admitted at two centres in the North East of England over 3 years revealed 85 patients. Patients were divided into two groups depending on whether they were taking clopidogrel alone (C) or with aspirin (CA). Haemoglobin drop was significantly different in the CA group that was operated on early (CA1) versus the group for which surgery was delayed by over 48 hours (CA2): 3.3g/dl and 1.9g/dl respectively (p=0.01). The mean inpatient stay in group C was 35.9 days while in group CA it was 19.9 days (p=0.002). The mean length of stay in group CA2 (26.7 days) was significantly longer than for CA1 patients (14.1 days) (p=0.01). There were no significant differences in mortality or wound complications. Hip fracture patients on clopidogrel can be safely operated on early provided they are medically stable. Bleeding risk should be borne in mind in those patients on dual therapy with aspirin.

The Best Practice Tariff helps improve management of neck of femur fractures: a completed audit loop.

BACKGROUND: The Best Practice Tariff incentivizes hospitals in the UK to improve the care they deliver, and includes a requirement to deliver multiprofessional care to patients with neck of femur fractures. The Best Practice Tariff for 2010-11 included six targets: (1) surgery within 36 hours, (2) admission under consultant-led joint orthopaedic-geriatric care, (3) admission using a multidisciplinary assessment protocol, (4) review by a geriatrician within 72 hours, (5) geriatrician-directed multi-professional rehabilitation, and (6) assessment for falls and bone protection. The authors chose to audit their Trust's compliance with these targets. METHODS: A retrospective audit was conducted in 2011 at the authors' university-affiliated tertiary care hospital, which is a regional major trauma centre. Only patients 65 years or older, with fragility-type neck of femur fractures who were treated surgically at the authors' unit and were eligible for geriatric review and multiprofessional rehabilitation, were included. The results of this audit (2010-11 Best Practice Tariff targets) were analysed and a series of procedural and logistical measures were introduced. A re-audit was performed in April 2012 for 2011-12, and the results for the 2 years were compared using appropriate statistics (Chi square tests and analysis of variance). Thirty-day mortality was compared using the summary hospital-level mortality indicator. RESULTS: A total of 410 patients were eligible for Best Practice Tariff in 2010-11, which increased to 463 in 2011-12. The changes from the first year's audit helped increase the rates for 36-hour surgery from 48.3% to 73.4% and for 72-hour geriatric review from 68.8% to 81.8% (P<0.05). The annual Best Practice Tariff achievement increased from 31.7% to 61.3% (P<0.05). The summary hospital-level mortality indicator declined from 96.5 to 61.3. CONCLUSIONS: Focusing on poorly satisfied Best Practice Tariff indicators can produce a significant improvement in the per capita Best Practice Tariff achievement. Further studies are needed to assess the health and financial gain in detail.

Sequential expression of VEGF and its receptors in human placental villi during very early pregnancy: differences between placental vasculogenesis and angiogenesis.

Vascularization within the human placenta is the result of the de novo formation of vessels derived from pluripotent precursor cells in the mesenchymal core of the villi. Vascularization of placental villi starts at around day 21 post conception (p.c.) with a four somite embryo. At this stage progenitors of haemangiogenic cells differentiate to form first vessels. These progenitor cells are thought to be directly derived from mesenchymal cells rather than originating from fetal blood cells. We investigated the relation between differentiation of stromal cells towards endothelial cells and vascular structures and the expression pattern of the respective growth factors. Using transmission electron microscopy and immunohistochemistry (for VEGF, Flt-1, Flk-1, CD14, CD34, and CD68) the development of placental vasculogenesis during very early stages of pregnancy (days 22-48 p.c.) was studied. We found that VEGF is strongly expressed in villous cytotrophoblast cells and subsequently in Hofbauer cells while its receptors Flt-1 and Flk-1 are found on vasculogenic and angiogenic precursor cells. The developmental expression and secretion of VEGF suggests its involvement in recruitment, maintenance and formation of first angiogenic cells and vessels. Interactions between VEGF and Flk-1 and Flt-1 may regulate placental vasculogenesis and angiogenesis in a paracrine and autocrine manner. The sequential expression of growth factors in different cell types may point to the fact that placental vasculogenesis and angiogenesis are clearly distinct events.

Structural differentiation of human uterine luminal and glandular epithelium during early pregnancy: an ultrastructural and immunohistochemical study.

The differentiation of human endometrial epithelium is a dynamic event that occurs throughout the menstrual cycle and early pregnancy. The structural transformation and differentiation of human uterine luminal and glandular epithelium of early human pregnancy (n=14) was investigated ultrastructurally and immunohistochemically using antibodies against cytokeratin (CT), endothelial marker CD31, Fas, and proliferating cell nuclear antigen (PCNA). Ultrastructurally, luminal epithelial cells showed distinctive euchromatic nuclei with prominent nucleoli and relatively loose cell membranes in all poles (apical to basal). Subcellular components were easily recognized in luminal epithelium except in degenerating cells. Mainly two cell types, dark and clear cells, formed the glandular epithelium. In the early gestation period, microvilli were abundant on the apical and apico-lateral poles of these cells. Only a few cytoplasmic projections were observed in dark cells. Numerous cilia were observed on the apical pole of some clear cells, located at the adluminal segment. In contrast, dark cells lacked cilia, nuclear channels, or giant mitochondrial profiles. Glycogen synthesis and apocrine secretion were recognizable for several days during early gestation. The apocrine secretory activity differed among dark cells of the glandular epithelium. The immunoreactivity of PCNA and Fas, and ultrastructural observations in the glandular epithelium suggest that, even in different segments of the same gland, epithelial cells do not regress during early gestation, but proliferate, perhaps representing a resistance against trophoblastic invasion. These morphological and molecular changes suggest that both luminal and glandular epithelium may play an important role in cellular defense and limitation for trophoblastic invasion during early pregnancy since plasma membrane alterations of the surface epithelium take place at the apical, basal and lateral poles compared to early secretory phase endometrial cells. Besides glandular epithelium may be consequently responsible for uterine secretions, which may be critical for early embryo development.

Body composition, bone mass and microstructural analysis in GH-transgenic mice reveals that skeletal changes are specific to bone compartment and gender.

Experimental and clinical studies suggest that high serum levels of growth hormone (GH) increase cortical but not trabecular bone. We studied body composition and bone structure in transgenic mice (MT-bGH) with systemic overexpression of GH. Body composition was examined with dual-energy X-ray absorptiometry (DXA), ashing, and chemical analysis, and the femora with DXA and micro computerized tomography. The absolute fat and bone tissue contents were significantly higher in GH transgenic mice vs controls (P < or = 0.05), but no significant difference was noted when normalizing the values to body weight. Male transgenics displayed no change in apparent (volumetric) femoral bone density, relative cortical area and trabecular bone volume fraction. Female transgenic mice demonstrated an increase in apparent femoral density and in trabecular bone volume fraction (+130%; P < or = 0.01). The mineralized tissue matrix density was decreased in male and female transgenic mice (P < or = 0.05). The results show that chronic GH excess affects trabecular bone in a gender-specific manner and that bone changes depend on the compartment investigated.

Precision of high-resolution dual energy X-ray absorptiometry of bone mineral status and body composition in small animal models.

The purpose of this study was to analyze the in situ precision (reproducibility) of bone mineral and body composition measurements in mice of different body weights and rats, using a high-resolution DXA (dual energy X-ray absorptiometry) scanner. We examined 48 NMRI mice weighing approximately 10 to 60 g, and 10 rats weighing approximately 140 g. Four repeated measurements were obtained on different days. In mice, the standard deviations of repeated measurements ranged from 2.5 to 242 mg for bone mineral content (BMC), from 0.16 to 3.74 g for fat, and from 0.40 to 4.21 g for lean mass. The coefficient of variation in percent (CV%) for BMC/BMD (bone mineral density) was highest in the 10 g mice (12.8% / 4.9%) and lowest in the 40 g mice (3.5% /1.7%). In rats, it was 2.5 /1.2% in the lower extremity, 7.1/3.0 % in the spine, 5.7/2.0 % in the femur, and 3.6%/2.1% in the tibia. The CV% for fat and lean mass in mice was higher than for BMC. The study demonstrates good precision of bone mineral and moderate precision of body composition measurements in small animals, using a high-resolution DXA system. The technique can be used for testing the efficacy of drugs in small animal models, for mutagenesis screens, and for the phenotypic characterization of transgenic mice.

Menstruum induces changes in mesothelial cell morphology.

In previous studies, we have shown that menstrual endometrium preferentially adheres to the subepithelial lining of the peritoneum. It remains to be elucidated, however, whether this damage is preexisting or inflicted by the menstrual tissue itself. We hypothesized that the menstrual tissue itself damages the peritoneum. To investigate this, the viability of menstrual endometrial tissue in peritoneal fluid (PF) was evaluated and the morphologic changes in the mesothelial cells were studied by in vitro cocultures of menstruum with mesothelial cell monolayers. Menstruum was collected with a menstrual cup. Endometrial tissue was isolated from the menstruum, resuspended in culture medium or in the cell-free fraction of PF and cultured for 24, 48 or 72 h. A 3(4, 5-dimethylthiazolyl-2)-2,5-diphenyl tetrazolium bromide (MTT) assay was performed to obtain a relative measure of viable adhered endometrial cells. Mesothelial cells isolated from human omental tissue were cultured on Matrigel or uncoated plastic. At confluence, overnight cocultures were performed and scanning electron microscopy was used to evaluate the morphologic changes. The viability of endometrial fragments was 84% (n = 36, p < 0.05), 82% (n = 27, not significant) and 104% (n = 14, not significant) when cultured in the cell-free fraction of PF for 24, 48 and 72 h, respectively, when compared to medium with 10% fetal calf serum. Menstrual endometrial fragments or menstrual serum added to and cocultured with mesothelial cells induced severe morphologic alterations of the latter, including retraction, shrinking and gap formation. Similar morphologic changes were observed when mesothelial cells were cocultured with menstrual endometrial fragments in PF or in culture inserts. Incubation with conditioned medium from cultured menstrual endometrium induced similar but less pronounced changes in morphology. In conclusion, menstrual endometrial fragments remain viable in PF in vitro for at least 72 h. Antegradely shed menstruum induces changes in mesothelial cell morphology, including retraction and shrinking with exposure of the underlying surface. These findings suggest that menstruum is harmful to the peritoneal lining. Therefore, by local destruction of the mesothelial layer, menstrual endometrium is able to create sites for adhesion.

Morphological changes in mesothelial cells induced by shed menstrual endometrium in vitro are not primarily due to apoptosis or necrosis.

In a previous study on the pathogenesis of endometriosis, we observed that constituents of menstrual effluent induce morphological alterations in human mesothelial cells. In this study, we investigated whether these alterations were associated with apoptosis or necrosis or were the result of cellular remodelling. After overnight incubation of confluent monolayers of human omental mesothelial cells (HOMEC) with conditioned media prepared from menstrual effluent shed anterogradely, severe alterations in morphology were observed. Typical polygonal mesothelial cell cultures at confluency acquired elongated spindle morphology, resulting in gaps between the cells. In contrast, mesothelial cells from the control groups receiving culture medium only, retained a normal morphology. Immunofluorescence staining revealed that cytokeratin, vimentin and actin filaments were still present, homogeneously distributed in the cell cytoplasm following changes in morphology. To evaluate whether the morphological alterations were associated with apoptosis and/or necrosis, the cells were stained with the M30 CytoDeath antibody or annexin V with propidium iodide and analysed using flow cytometry. The results showed that only a small percentage (1-7%) of the affected HOMEC were undergoing apoptosis or necrosis. We conclude that the profoundly altered morphology of HOMEC is a result of cellular remodelling and that the role of apoptosis and necrosis is negligible. Soluble paracrine factors released by cells isolated from menstrual effluent shed anterogradely may induce a reorganization of the cytoskeleton. As a result, the underlying basement membrane will be exposed and the mesothelium may no longer prevent implantation of endometrium shed retrogradely into the peritoneum, thus facilitating the development of endometriosis.

[Sex-specific analysis of bone mass in normal and growth hormone transgenic mice using dual energy x-ray absorptiometry (DXA)]. / Geschlechtsspezifische Analyse der Knochenmasse normaler und Wachstumshormon-transgener Mäuse mittels Zweienergie-Röntgen-Absorptiometrie (DXA).

The aim of the present study was the non-invasive, sex-specific measurement of bone mass in bovine growth hormone (bGH) transgenic mice and normal controls with dual energy X-ray absorptiometry (DXA). The transgenic mouse constitutes a suitable animal model to study the influence of growth hormone on the skeletal system. We analysed 28 animals, aged 12 weeks (14 transgenic, 14 controls, 7 male and 7 female, respectively), using a peripheral DXA scanner that had been adapted to the measurement of small animals. At a measurement time of 20 min, the precision (RMS average CV%) was 4.4% for bone mass (BMC), 2.5% for areal bone density (BMD), 0.86% for total body weight and 4.5% for the percentage BMC (relative to body weight). While the absolute bone mass was not significantly different between male and female animals, we found a higher percentage of the BMC relative to the total bone mass in females (+21% in controls, +31% in transgenics; p < 0.01). The absolute bone mass was higher in the transgenic animals (+71% in females and +62% in males; p < 0.01), but relative to the body weight the transgenic females yielded similar and the transgenic males lower values (-7.2%; p < 0.05). Using DXA it is possible to non-invasively determine the mass of mineralised tissue in the mouse with relatively high precision and to effectively discriminate between different groups. Although a strong influence of growth hormone on the absolute bone mass is observed, the results show that this increase is not higher than that of the total body mass.