The power of the Mediator complex-Expanding the genetic architecture and phenotypic spectrum of MED12-related disorders.

MED12 is a member of the large Mediator complex that controls cell growth, development, and differentiation. Mutations in MED12 disrupt neuronal gene expression and lead to at least three distinct X-linked intellectual disability syndromes (FG, Lujan-Fryns, and Ohdo). Here, we describe six families with missense variants in MED12 (p.(Arg815Gln), p.(Val954Gly), p.(Glu1091Lys), p.(Arg1295Cys), p.(Pro1371Ser), and p.(Arg1148His), the latter being first reported in affected females) associated with a continuum of symptoms rather than distinct syndromes. The variants expanded the genetic architecture and phenotypic spectrum of MED12-related disorders. New clinical symptoms included brachycephaly, anteverted nares, bulbous nasal tip, prognathism, deep set eyes, and single palmar crease. We showed that MED12 variants, initially implicated in X-linked recessive disorders in males, may predict a potential risk for phenotypic expression in females, with no correlation of the X chromosome inactivation pattern in blood cells. Molecular modeling (Yasara Structure) performed to model the functional effects of the variants strongly supported the pathogenic character of the variants examined. We showed that molecular modeling is a useful method for in silico testing of the potential functional effects of MED12 variants and thus can be a valuable addition to the interpretation of the clinical and genetic findings.

A Novel Approach for Non-Invasive Continuous In-Line Control of Perfusion Cell Cultivations by Raman Spectroscopy.

Continuous manufacturing is becoming more important in the biopharmaceutical industry. This processing strategy is favorable, as it is more efficient, flexible, and has the potential to produce higher and more consistent product quality. At the same time, it faces some challenges, especially in cell culture. As a steady state has to be maintained over a prolonged time, it is unavoidable to implement advanced process analytical technologies to control the relevant process parameters in a fast and precise manner. One such analytical technology is Raman spectroscopy, which has proven its advantages for process monitoring and control mostly in (fed-) batch cultivations. In this study, an in-line flow cell for Raman spectroscopy is included in the cell-free harvest stream of a perfusion process. Quantitative models for glucose and lactate were generated based on five cultivations originating from varying bioreactor scales. After successfully validating the glucose model (Root Mean Square Error of Prediction (RMSEP) of ∼0.2 g/L), it was employed for control of an external glucose feed in cultivation with a glucose-free perfusion medium. The generated model was successfully applied to perform process control at 4 g/L and 1.5 g/L glucose over several days, respectively, with variability of ±0.4 g/L. The results demonstrate the high potential of Raman spectroscopy for advanced process monitoring and control of a perfusion process with a bioreactor and scale-independent measurement method.

Aprotinin-associated risks in off-pump coronary artery bypass grafting.

BACKGROUND: Little data is available regarding the safety of using the serine protease inhibitor aprotinin in off-pump cardiac surgery. We retrospectively assessed the risks of administering the drug to adult patients undergoing off-pump coronary artery bypass grafting (OPCABG). METHODS: Aprotinin was administered as a bolus of 1 or 2 million kallikrein inhibiting units to 391 patients following median sternotomy; 370 control patients underwent surgery during the same time period without receiving aprotinin. No other antifibrinolytic agents were administered. RESULTS: Preoperative characteristics, length of ICU and hospital stay were similar between the mostly medium-risk aprotinin and the control patients. Postoperative cardiac, renal, neurological, and respiratory complications and hospital mortality occurred with comparable frequencies in both groups. Levels of myocardial enzymes during the first 72 h after surgery also did not differ significantly. CONCLUSION: Use of aprotinin in OPCABG was not associated with a higher incidence of hospital mortality, cardiovascular, renal, or other complications. Given the good safety profile in this large patient population we suggest that aprotinin could still be a valid antifibrinolytic treatment option in OPCABG.

DNA degradation during maturation of erythrocytes - molecular cytogenetic characterization of Howell-Jolly bodies.

Howell-Jolly bodies (HJBs) are small DNA-containing inclusions of erythrocytes and are often present after splenectomy. The genetic composition of HJBs is unknown at present. We isolated individual erythrocytes that had inclusion bodies from five splenectomized patients and performed DNA amplification using degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR) with subsequent reverse painting on normal male metaphase spreads. We also measured the sizes of HJBs in erythrocytes from a splenectomized patient using an inverted microscope. Two-dimensional positions of HJBs were projected onto a virtual erythrocyte. The average size of HJBs was 0.73 +/- 0.17 microm (range 0.4-1.1 microm). Inside the erythrocyte the HJBs were found to be equally distributed. Small HJBs contained DNA from one or two centromeres and larger HJBs contained DNA from up to eight different centromeres. Centromeric DNA from chromosomes 1/5, 7, 8, and 18 was most frequently observed. Signals from the centromeric regions of chromosomes 3, 4, 9, and 10 were not observed. Signals from euchromatic regions were detected in a few cases. We hypothesize that in addition to enucleation and nucleus fragmentation DNA degradation during maturation of erythrocytes preferentially eliminates euchromatic DNA. Similarities between these processes and those described for embryonic stem cells suggest that most stem cells are able to degrade DNA in a genetically controlled manner.

Onto better TRAILs for cancer treatment.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), also known as Apo-2 ligand (Apo2L), is a member of the TNF cytokine superfamily. By cross-linking TRAIL-Receptor (TRAIL-R) 1 or TRAIL-R2, also known as death receptors 4 and 5 (DR4 and DR5), TRAIL has the capability to induce apoptosis in a wide variety of tumor cells while sparing vital normal cells. The discovery of this unique property among TNF superfamily members laid the foundation for testing the clinical potential of TRAIL-R-targeting therapies in the cancer clinic. To date, two of these therapeutic strategies have been tested clinically: (i) recombinant human TRAIL and (ii) antibodies directed against TRAIL-R1 or TRAIL-R2. Unfortunately, however, these TRAIL-R agonists have basically failed as most human tumors are resistant to apoptosis induction by them. It recently emerged that this is largely due to the poor agonistic activity of these agents. Consequently, novel TRAIL-R-targeting agents with increased bioactivity are currently being developed with the aim of rendering TRAIL-based therapies more active. This review summarizes these second-generation novel formulations of TRAIL and other TRAIL-R agonists, which exhibit enhanced cytotoxic capacity toward cancer cells, thereby providing the potential of being more effective when applied clinically than first-generation TRAIL-R agonists.

A comparative trial of imipenem versus ceftazidime in the release of endotoxin and cytokine generation in patients with gram-negative urosepsis. Urosepsis Study Group.

Evidence from in vitro experiments and animal and human studies indicate that antibiotic therapy may induce the release of endotoxin from the outer membrane of Gram-negative bacteria. Antibiotics that bind preferentially to penicillin-binding protein-2 (PBP-2)--such as imipenem--are associated with little release of endotoxin, while antibiotics that preferentially bind to PBP-3--such as ceftazidime--are associated with far greater release of endotoxin. We conducted a randomized, multicenter, double-blind study comparing imipenem to ceftazidime in patients with urinary tract infections caused by Gram-negative bacilli associated with signs and symptoms of systemic inflammation. A total of 33 patients were randomized to receive either imipenem (n = 14) or ceftazidime (n = 19) for initial treatment for urosepsis. No differences in plasma endotoxin, interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha) or urine endotoxin, IL-6 or IL-8 levels were found between the two treatment groups within the first 8 h after antibiotic administration. We conclude that, if differences exist with respect to endotoxin release by these two antimicrobial agents, these differences are not readily demonstrable in this clinical study with carefully defined patients with Gram-negative urinary tract infections.

The relationship of bone mass and fracture history to fluoride and calcium intake: a study of three communities.

Stimulated by the suggestion that water fluoride greater than 1 mg/L may protect against osteoporosis, we studied bone mass of women in three rural communities with differing mineral content of the water supply. Mean fluoride and calcium of community drinking waters were 4 mg/L and 16 mg/L, respectively, high fluoride community; 1 mg/L and 375 mg/L, respectively, high calcium community; and 1 mg/L and 65 mg/L, respectively, low calcium community. Bone mass was measured by single photon absorptiometry, and women were interviewed about fracture history, dietary intake, and other important covariates. We observed no protective effect with higher fluoride intake. Bone mass was lower in older women from the high fluoride community though not statistically so; these women reported significantly more fractures. There was no observed community difference in young women's bone mass or fracture history. Young women in the high fluoride community consuming calcium and vitamin D in excess of 800 mg/day and 400 IU/day, respectively, had significantly better bone mass (p less than 0.05) than their peers.

Correlates of mid-radius bone density among postmenopausal women: a community study.

We surveyed mid-radius bone density in a geographically-defined population of 324 women, 55-80 years old from two small, demographically-similar communities whose water supply provided 60 mg/l and 375 mg/l of elemental calcium. Mean community calcium intakes were 964 mg/day and 1329 mg/day respectively. Bone density, measured by photon densitometry, was correlated with physical measurements, medical history, and reported nutritional intake. Mid-radius bone density decreased with age while the following additional factors were independently and positively associated with bone density: humeral muscle area (p = 0.0001), extended estrogen use (p = 0.0004), thiazide use (p = 0.0029), and vitamin D intake (p = 0.0104). Estimated total calcium intake alone did not significantly correlate with bone density; however, mean bone density was significantly greater in persons whose calcium intake was greater than 800 mg/day, consumed concurrently with vitamin D in amounts greater than 400 IU (p = 0.0342), the Recommended Daily Allowance of these two nutrients.

The association of intakes of vitamin D and calcium with blood pressure among women.

Blood pressure and its relationship to nutritional factors, particularly intakes of calcium and vitamin D, were evaluated among 86 women, aged 20-35 yr, and 222 women, aged 55-80 yr, who were not using diuretics. Observations were based on a study of bone density among women in two communities which included blood pressure determinations, a food frequency estimate of calcium intake, a 24-h dietary recall and an extensive supplement use history. There was no significant relationship between estimated current dietary calcium consumption and blood pressure in either age group. However, in younger women, there was a significant inverse relationship between estimated dietary intake of vitamin D and systolic blood pressure which remained significant (p = 0.0016) after adjusting for age, Quetelet index, alcohol consumption and calcium intake. Older women whose consumption of both vitamin D and calcium was less than the Recommended Dietary Allowance, 400 IU/day and 800 mg/day respectively, had a significantly higher systolic blood pressure (p = 0.0371) than their counterparts whose estimated intake met the RDA for at least one of the two nutrients. Vitamin D may be related to blood pressure through its regulation of calcium absorption from the gut or its interaction with parathyroid hormone in maintaining plasma calcium homeostasis.

Parameters related to 25-OH-D levels in a population-based study of women.

We surveyed calcidiol levels (25-OH-D) in a geographically-defined population of 373 women, ages 20-80 yr to test for an association between occult osteomalacia and excess bone loss. Bone mass was measured by photon densitometry and an estimate of vitamin D was determined by measuring dietary and supplemental intake as well as sunlight exposure equivalent. The relationship of smoking practices, alcohol use, exogenous estrogen use, and medications to calcidiol level was assessed. Calcidiol levels were not associated with bone mass levels observed at two different forearm sites. Low levels of calcidiol, indicative of generalized vitamin D deficiency, were not observed in this population though mean estimates of vitamin D intakes from food 119 +/- 148 IU) or food and supplements (319 +/- 463 IU) were less than the Recommended Dietary Allowance (400 IU). Calcidiol was significantly associated with estimates of vitamin D intake from food (r = 0.11), supplement use (r = 0.21), and sunlight equivalent exposure (r = 0.26). Calcidiol levels were negatively related to age (p = 0.0020) and positively related to exogenous estrogen use and premenopausal state independent of age.

Relationship between 1,25-dihydroxyvitamin D and blood pressure in a geographically defined population.

In a cross-sectional, population-based study we measured casual, seated blood pressure with a random-zero sphygmomanometer and 1,25-dihydroxyvitamin D (1,25-[OH]2D) with a protein-binding assay in 373 women aged 20-80 y. 1,25-(OH)2D, an active metabolite that regulates serum calcium, was associated significantly and positively with systolic blood pressure (p = 0.020) and diastolic blood pressure (p = 0.003) after adjustment for age, Quetelet's index (a measure of obesity), and current thiazide use. A model including age, Quetelet's index, current thiazide use, and 1,25-(OH)2D explained 37% of the variability in systolic blood pressure observations, of which 7% of variability was explained by 1,25-(OH)2D. In this geographically defined population of women, the variability of blood-pressure measurements attributable to 1,25-(OH)2D was of the same order of magnitude as that attributable to Quetelet's index.

Divalent ion handling in human kidney donors with increased blood pressure after uninephrectomy.

In a prospective study of 21 normal human kidney donors, increases in blood pressure were found in seven of 12 males and in three of nine females within the first year after uninephrectomy. Donors with blood pressure increases were characterized by greater weight and body mass indexes. In addition, urinary phosphate excretion was positively correlated (r = 0.73, p less than 0.001) and tubular reabsorption of phosphate negatively correlated (r = -0.61, p less than 0.01) with blood pressure at the follow-up periods in which increases were observed. All donors experienced an increase in parathyroid hormone levels and urinary cyclic AMP excretion. These changes were accompanied by decreases in urinary calcium and serum phosphate values. Thus, the increase in blood pressure took place in a milieu similar to that of "normocalcemic" hyperparathyroidism. The correlation of phosphate excretion and blood pressure in normal donors suggests an important role for phosphate metabolism in the genesis of hypertension associated with loss of renal mass.

Age-related hearing loss and bone mass in a population of rural women aged 60 to 85 years.

Demineralization of the cochlear capsule in conjunction with age-related bone mass loss may be one biologic factor contributing to hearing loss in the elderly. In other metabolic bone diseases, including Paget's disease of the bone and cochlear otosclerosis, demineralization of the cochlea has been associated with sensorineural hearing loss. In 1988/1989, the relation between hearing loss and bone mass of the radius and femoral neck was studied cross-sectionally in 369 women aged 60 to 85 years from three rural communities. Hearing sensitivity was measured using audiometry, and bone mineral density of the radius and femoral neck was measured using single- and dual-photon densitometry, respectively. Three variables, ascertained by interview, were associated with an increased odds for hearing loss: age, family history of hearing loss before the age of 50 years, and current use of more than two nonestrogen, nonthiazide prescription medications. After consideration of the effect of these three variables, women with femoral neck bone mass values below the mean value of 0.696 g/cm2 for this population had a 1.9 greater odds of having a hearing loss (confidence interval: 1.30, 2.50). This study demonstrated a consistent adjusted association between femoral neck bone mass and age-related hearing loss in a population of rural women aged 60 to 85 years. No consistent association was observed between radial bone mass and hearing loss.

Statistical analysis of K 2 x 2 tables: a comparative study of estimators/test statistics for association and homogeneity.

In order to control for confounding variables, epidemiologists often obtain data in the form of a 2 x 2 table. One variable is usually the disease status, while the other variable represents a dichotomous exposure variable that is suspected of being a risk factor. If a confounding variable is present, the data are often stratified into several 2 x 2 tables. The objectives of the analysis are to test for the association between the suspected risk factor and the disease and to estimate the strength of this relationship. Before estimating a common odds ratio, it is important to check whether the odds ratios are homogeneous. This paper presents the results of a Monte Carlo study that was performed to determine the size and power of a number of tests of association and homogeneity when the data are sparse. We also evaluated the performance of three estimators of the common odds ratio. For the Monte Carlo studies, equal numbers of cases and controls were used in a wide variety of sparse data situations. On the basis of these studies, we recommend the Breslow-Day test for nonsparse data, and the T4 and T5 statistics for sparse data to test for homogeneity. The Mantel-Haenszel test of association is recommended for sparse and nonsparse data sets. With sparse data, none of the odds ratio estimators are entirely satisfactory.

De novo complete trisomy 5p: clinical report and FISH studies.

We describe a de novo trisomy 5p in a 1-year-old severely retarded boy. The complete short arm of chromosome 5 segregated as an additional marker chromosome in all metaphases. The marker was identified as 5p by conventional cytogenetic techniques (GTG, GBG, CBG) and molecular cytogenetic techniques (whole chromosome-painting probe, probes for the cri-du-chat region and the centromere, and additionally high-resolution multicolor banding using a chromosome 5-specific DNA probe cocktail). The clinical findings were similar to the established trisomy 5p phenotype including macrocephaly, facial abnormalities, tracheobronchial defects with subsequent respiratory infections, hypotonia, and psychomotor retardation. To the best of our knowledge this is the first description of an isolated complete 5p trisomy without involvement of the aberrant chromosome in any structural chromosomal rearrangements.

Urinary incontinence in rural older women: prevalence, incidence and remission.

OBJECTIVES: To determine the prevalence, incidence, and remission rates of urinary incontinence in a large group of older women over a 6-year time span and to assess factors associated with incontinence incidence and remission. DESIGN: Longitudinal cohort study. SETTING: Two rural counties in Iowa. PARTICIPANTS: 2025 women aged 65 years or older residing in rural Iowa, enrolled in the Iowa 65+ Rural Health Study of EPESE (Establishment of Populations for Epidemiologic Studies of the Elderly) were interviewed in person annually for 6 years; specific responses to queries about urinary incontinence were given at baseline, 3-, and 6-year intervals. MEASUREMENTS: Conditional multivariate logistic regression analysis was done to assess the relationship between incontinence symptoms and various factors previously found to be related to incontinence. RESULTS: The baseline prevalence of urge incontinence was 36.3%, and of stress incontinence it was 40.3%. For urge incontinence, the 3-year incidence and remission rates between the third and sixth years were 28.5% and 22.1%, respectively. For stress incontinence, the 3-year incidence and remission rates between years 3 and 6 were 28.6% and 25.1%, respectively. Seventy-six percent and 84% of women who reported no urge or stress incontinence, respectively, at the baseline interview were continent at both follow-up interviews. The only significant factors related to changes in incontinence status were age, which was associated with an increased incidence of urge incontinence (OR 1.11, P = .017, 95% CI 1.019-1.203), and improvement in activities of daily living, which was associated with a increased remission of urge incontinence (OR 0.50, P = .015, 95% CI 0.28-0.9) CONCLUSION: In some older women, urinary incontinence is a dynamic state, with women moving back and forth along a continuum between continence and incontinence. These results are tempered by limitations of the study, which include its questionnaire design and lack of ability to detect potential treatment effect.

An introduction to time-trend analysis.

Healthcare professionals often are presented with data that appear to indicate an upward or downward trend over time. For example, admissions of acquired immunodeficiency syndrome (AIDS) patients appear to be increasing, cesarean section rates appear to be decreasing, or nosocomial pneumonia rates appear to be increasing. Critical decisions sometimes are based on such trends, which often are presented without a statistical analysis. Those responsible for decision making may be left wondering whether these apparent trends represent only chance variation. Graphs showing trends over time generally present one of three kinds of outcome data: counts (eg, three AIDS admissions), proportions (eg, 10 cesarean sections per 100 total deliveries), or person-time data (eg, 13 cases of nosocomial pneumonia per 10,000 patient days). Using familiar examples and a minimum of technical language, we illustrate the analysis of time trends.

Prevalence and characteristics of multiple analgesic drug use in an elderly study group.

With few exceptions, use of multiple analgesic drugs achieves dubious increases in analgesia while placing elders at increased risk of the many potential adverse effects of analgesic drugs. The potential for duplication of analgesic therapy among the elderly is great due to prevalent painful chronic conditions and the variety of prescription and nonprescription analgesic remedies available. The prevalence of multiple analgesic product use and patterns of concurrent use of different analgesic categories was investigated in a geographically defined population of persons 65 years of age and older. The demographic characteristics of users of multiple analgesic drug products were examined, as were their smoking status, alcohol use, lifetime history rates of major illnesses, physical functioning, pain experiences, memory performance, and depressive symptoms. A substantial proportion of analgesic users reported taking multiple products in the preceding 2 weeks (14.4% of female and 10.5% of male analgesic users). Men who reported pain in the preceding year were more likely to use multiple analgesic products. Women who experienced pain or limited physical functioning, or who had higher depressive symptom scores or a life-time history of ulcers were most likely to use multiple analgesic products. Thus, although some users of multiple analgesic products reported significant pain, several other factors were shown to be related to the phenomenon of multiple use.

Prevalence and functional correlates of low back pain in the elderly: the Iowa 65+ Rural Health Study.

As part of an interview survey of a defined population of 3,097 rural persons 65 years and older (the Iowa 65 + Rural Health Study), the prevalence and functional correlates of specifically defined low back pain were studied. Low back pain was reported by 23.6 per cent of the women and 18.4 per cent of the men in the year prior to the survey, with prevalence rates declining with age. Forty per cent of those with low back pain noted its presence at the time of the interview. Over half had used analgesics. Use of medical and chiropractic services for this symptom was nearly 75 per cent; 25 per cent had at least one hospitalization directly related to low back pain and over 5 per cent had low back surgery. Limitation of walking, sitting, bending over, and performing household chores was reported by 15 to 40 per cent, and 21 per cent attributed sleep disturbance to the low back pain. Nearly 75 per cent of subjects with low back pain reported first onset of the problem prior to age 65, which has important implications for pathogenesis and prevention. The functional and clinical burdens of low back pain in this population of rural elderly were substantial and require further evaluation of risk factors and impact on health status.

Medication use characteristics in the elderly: the Iowa 65+ Rural Health Study.

Medication use was studied in a rural, elderly population. Household interviews were conducted of 3,467 individuals aged 65 years or older. A total of 9,955 prescription or nonprescription drugs were reported by the respondents. The overall mean number of drugs per respondent was 2.87, while 12% of all respondents were not taking any drugs. Mean prescription and overall drug use increased significantly with increasing age (P less than .001), while mean nonprescription drug use was relatively constant across age groups. Significantly more women were prescription and nonprescription drug users. Directions for scheduled daily dosing accounted for 75% of all directions. The majority of prescription and nonprescription drugs had been taken on the previous day. General practitioners accounted for more prescription drugs (39.7%) than any other medical specialty. The most frequently stated purpose was cardiovascular for prescription drugs and musculoskeletal for nonprescription drugs. The three most frequent prescription drug therapeutic categories were cardiovascular (54.7%), central nervous system (CNS) agents (11.4%), and analgesics (9.4%). For nonprescription drugs, the three most frequent therapeutic categories were analgesics (39.6%), vitamins and minerals (32.9%), and laxatives (14.1%). Implications of these findings are discussed.