Hoechst staining and exposure to UV laser during flow cytometric sorting does not affect the frequency of detected endogenous DNA nicks in abnormal and normal human spermatozoa

Controlling the sex of offspring by the separation of X and Y chromosome-bearing spermatozoa using flow cytometry has been reported as a clinical technique aiding prevention of X-linked diseases. Although this technique has resulted in several hundred normal births in animals and at least one human birth, there is still concern over its genetic safety due to the involvement of two potentially mutagenic agents: UV light and the fluorochrome dye, Hoechst 33342 (H33342). Human spermatozoa, particularly those considered abnormal, may be more likely to suffer DNA damage following exposure to mutagenic agents, compared with other mammalian species. The stability of normal fresh and decondensed human spermatozoa were examined after exposure to a range of levels of UV and H33342 staining, using an assay that detects endogenous nicks in the DNA of spermatozoa. The stability of abnormal and normal, fresh and frozen-thawed human spermatozoa was examined following UV laser, H33342 staining and flow cytometry treatments utilizing the same assay. There was an increase in the presence of endogenous nicks when spermatozoa were decondensed compared with fresh spermatozoa. There was no increase in the incidence of nicks in any group of spermatozoa after UV and fluorochrome exposure compared with controls without exposure.      

Modulation of disease severity of dystrophic epidermolysis bullosa by a splice site mutation in combination with a missense mutation in the COL7A1 gene

Dystrophic epidermolysis bullosa (EBD) is a clinically heterogeneous skin disorder, characterized by abnormal anchoring fibrils (AF) and loss of dermal-epidermal adherence. EBD has been linked to the COL7A1 gene at chromosome 3p21 which encodes collagen VII, the major component of the AF. Here we investigated two unrelated EBD families with different clinical phenotypes and novel combinations of recessive and dominant COL7A1 mutations. Both families shared the same recessive heterozygous 14 bp deletion at the exon-intron 115 boundary of the COL7A1 gene. The deletion caused in-frame skipping of exon 115 and the elimination of 29 amino acid residues from the pro-alpha1(VII) polypeptide chain. As a result, procollagen VII was not converted to collagen VII and the C-terminal NC-2 propeptide which is normally removed from the procollagen VII prior to formation of the anchoring fibrils was retained in the skin. All affected individuals also carried missense mutations in exon 73 of COL7A1 which lead to different glycine- to-arginine substitutions in the triple-helical domain of collagen VII. Combination of the deletion mutation with a G2009R substitution resulted in a mild phenotype. In contrast, combination of the deletion with a G2043R substitution led to a severe phenotype. The G2043R substitution was a de novo mutation which alone caused a mild phenotype. Thus, different combinations of dominant and recessive COL7A1 mutations can modulate disease activity of EBD and alter the clinical presentation of the patients.      

Infectious influenza A and B virus variants with long carboxyl terminal deletions in the NS1 polypeptides

An influenza A virus, A/turkey/Oregon/71, was shown by protein gel analysis to code for an NS1 protein approximately half the size of those of other influenza A viruses. Sequence analysis of the NS gene of this virus revealed a 10 nucleotide deletion resulting in an NS1 protein of only 124 amino acids. This truncated NS1 polypeptide retained its karyophilic pattern as detected by indirect immunofluorescence analysis of virus infected cells. Also, A/turkey/Oregon/71 virus grew to high titer in embryonated chicken eggs comparable to other influenza A viruses. We also identified a laboratory variant of an influenza B virus, clone 201, which codes for a truncated NS1 protein. Sequence analysis revealed a 13 nucleotide deletion resulting in a shortened NS1 protein of only 127 amino acids as compared to other influenza B virus NS1 proteins possessing a length of 281 amino acids. Again as shown for the NS1 proteins of other influenza B viruses the NS1 polypeptide of B virus clone 201 was found to localize in the nucleus of infected cells. It appears that large deletions in the carboxyl terminus of the NS1 proteins of influenza A and B viruses can be tolerated without affecting the functional integrity of the NS1 polypeptide.     

Bone island (enostosis): current concept — a review

An enostosis or bone island represents a focus of mature compact (cortical) bone within the cancellous bone (spongiosa). Thought by some to be a tumor-like condition and by others a hamartoma, this benign lesion is probably congenital or developmental in origin and reflects failure of resorption during endochondral ossification. A bone island can be virtually diagnosed based on its characteristic clinical and radiologic features. Typically asymptomatic, the lesion is usually an incidental finding, with a preference for the pelvis, femur, and other long bones, although it may be found anywhere in the skeleton, including the spine. Plain radiography reveals a homogeneously dense, sclerotic focus in the cancellous bone with distinctive radiating bony streaks (thorny radiation) that blend with the trabeculae of the host bone, creating a feathered or brush-like border. On CT scan, a bone island appears as a low-attenuation focus, and on MRI sequences it shows low signal intensity like cortical bone. A distinguishing feature of bone islands is that they are usually cold on skeletal scintigraphy. Thus, bone scan has been and continues to be the means of differentiating bone islands from the more aggressive entities. However, reports of histologically confirmed bone islands that were scintigraphically active have raised a note of caution about relying on this modality in the differential consideration of lesions otherwise characteristic of bone islands. Guides to the correct diagnosis should be looked for in the individual clinical situation and in the morphologic features of the lesion on plain radiography, CT, and MRI, without regard to the lesion's activity on bone scan. If such a lesion, however, is symptomatic and hot on scintigraphy, it demands close observation with follow-up imaging studies.     

Moderate alcohol consumption and bone density among postmenopausal women

Chronic alcohol abuse is associated with low bone density and high risk of fracture. However, moderate alcohol consumption may help to maintain bone density in postmenopausal women by increasing endogenous estrogens or by promoting secretion of calcitonin. We conducted a prospective study among a sample of 188 white postmenopausal women (ages 50-74) from the Nurses' Health Study who participated in a health examination between 1993 and 1995 that included bone density assessments of the lumbar spine and proximal femur. Long-term alcohol intake was calculated as the average of the 1980 and 1990 measures from a food frequency questionnaire. Women who consumed 75 g or more of alcohol per week had significantly higher bone densities at the lumbar spine compared with non-drinking women (0.951 vs. 0.849 g/cm2, p = 0.002) after adjusting for age, body mass index (kg/m2), age at menopause, use of postmenopausal estrogens, and smoking status. Further adjustment for physical activity and daily intakes of calcium, vitamin D, protein, and caffeine did not alter the results. We also observed a linear increase in spinal bone density over increasing categories of alcohol intake (p = 0.002), suggesting that alcohol intakes of less than 75 g/week may also be of benefit. This positive association was observed among both current users and never users of postmenopausal estrogens. In contrast to the lumbar spine, femoral bone density was not higher among drinkers compared with nondrinkers, although density did increase among drinkers with increasing level of alcohol consumption. Further research is needed to determine whether moderate alcohol consumption can help to protect against spinal fractures in postmenopausal women. This finding must also be evaluated within a larger scope of the risks and benefits of alcohol on heart disease, breast cancer, and hip fractures.     

Computerized bone densitometric analysis: operator-dependent errors

PURPOSE: To determine the nature and relative frequency of operator-dependent data analysis errors in dual x-ray absorptiometry. MATERIALS AND METHODS: Over 40 months, 2,528 dual x-ray absorptiometric examinations of the forearm, femoral neck, and lumbar spine were performed by 11 technologists by using standard techniques and software. Each analysis was reviewed by a radiologist; errors were recorded and corrected. RESULTS: There were no forearm analysis errors. There were 24 (0.9%) femoral neck analysis errors, of which 23 resulted from misplacement of the analysis region. There were 33 (1.3%) spinal analysis errors, of which 24 resulted from misplacement of intervertebral disk space markers. Analysis errors of the femur and spine resulted in six misdiagnoses (0.2%). CONCLUSION: Misdiagnosis due to analysis errors is rare. Femoral neck analysis errors were easily detectable, but accurate spinal analyses depended on accurate identification of vertebral end plates and posterior elements. Nonetheless, these potentially serious errors can be detected and corrected if the analyses are reviewed and interpreted by a supervising physician who is familiar with the relevant anatomy, proper analysis techniques, and factors--such as artifacts--that adversely affect the accuracy of the analysis.     

Sclerosing bone dysplasias — a target-site approach

Sclerosing bone dysplasias are a poorly understood group of developmental anomalies, much of whose etiology is still obscure. The list of conditions constituting this group is relatively short: osteopetrosis (Albers-Schönberg disease), pycnodysostosis (Maroteaux-Lamy disease), enostosis (bone island), osteopoikilosis, osteopathia striata (Voorhoeve disease), progressive diaphyseal dysplasia (Camurati-Engelmann disease), hereditary multiple diaphyseal sclerosis (Ribbing disease), four types of endosteal hyperostosis (van Buchem disease, Worth disease, Nakamura disease, and Truswell-Hansen disease), dysosteosclerosis, metaphyseal dysplasia (Pyle's disease), craniometaphyseal dysplasia, melorheostosis (Leri disease), and craniodiaphyseal dysplasia. There are instances in which two or more of the above disorders coexist. These are termed overlap syndromes, most commonly involving osteopathia striata, osteopoikilosis, and melorheostosis. A classification of these dysplasias is elaborated based on a target-site approach that views them as disturbances in development associated with the processes of either endochondral or intramembranous bone formation, or both. Accumulated evidence suggests that many of these disorders stem from common defects in bone resorption and/or formation during the processes of skeletal maturation and modeling. Finally, the subgroup of overlap syndromes is emphasized as indicating a strong interrelationship between the sclerosing dysplasias of bone, with perhaps a common pathogenesis for many.This article is one in a series of review articles which represent expansions of papers presented at the annual meeting of the International Skeletal Society and were solicited by the editors     

Use and need for post-acute services following paediatric head injury

This paper aims to document the types of inpatient and outpatient post-acute services children receive after discharge from an acute care hospital for head injury and to better understand the extent to which children fail to receive services and the reasons for not receiving needed services. A follow-up was conducted on 95 children (aged 5-15) 1 year after they were hospitalized for head injury. Parents were interviewed by phone concerning their child's use of and need for medical, rehabilitation, and social services since the injury. Questions were also asked regarding the child's current health status and behaviour. Inpatient records were reviewed to obtain information on the characteristics of the injury. Overall use of outpatient rehabilitation and social services was low during the year following injury, ranging from 0-18% of the study sample. Although need for and use of services was positively correlated with head injury severity, it appears that unmet need was highest for children with the least severe head injuries. Finally, need for physical or occupational therapy and mental health services was unrecognized for one third of children with physical limitations and 40% of children with at least 14 identified behaviour problems. These findings underscore the need for physicians and other health care professionals to thoroughly evaluate children during follow-up visits as well as during the initial hospitalization for head injury-related deficits. Identification of functional deficits or behavioural problems should be followed-up by evaluation and treatment by qualified rehabilitation professionals.     

Toxic cardiogenic shock in a patient receiving weekly 24-h infusion of high-dose 5-fluorouracil and leucovorin

A 54-year-old man was treated with weekly 24-h infusion of high-dose 5-fluorouracil (2600 mg/m2) and leucovorin (100 mg/m2) for metastatic colon cancer. At first, he tolerated the treatment well and no significant toxicity was identified. After a total of eight courses of treatment, a stable disease was observed, but mild shortness of breath was found on occasion. The patient had no previous history of cardiac disease and the heart performance assessed by left ventricular ejection fraction before treatment was normal. Unfortunately, acute pulmonary edema with lethal cardiogenic shock occurred during the ninth course of treatment, in spite of intensive medical treatment. The chest X-ray showed extreme cardiomegaly. Repeated assessment of his heart function by echocardiogram and ventricular ejection fraction revealed a very poor cardiac performance. Toxic cardiogenic shock during weekly 24-h infusion of high-dose 5-fluorouracil and leucovorin is extremely rare. To the best of our knowledge, no case has been reported in the English literature. We report a case and the relevant literature about the incidence, clinical picture and possible pathophysiology on 5-fluorouracil-related cardioxicity is reviewed.