RNA/DNA co-analysis from human skin and contact traces--results of a sixth collaborative EDNAP exercise.

The European DNA profiling group (EDNAP) organized a sixth collaborative exercise on RNA/DNA co-analysis for body fluid/tissue identification and STR profiling. The task was to identify skin samples/contact traces using specific RNA biomarkers and test three housekeeping genes for their suitability as reference genes. Eight stains, a skin RNA dilution series and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 22 participating laboratories using RNA extraction or RNA/DNA co-extraction methods. Two sets of previously described skin-specific markers were used: skin1 pentaplex (LCE1C, LCE1D, LCE2D, IL1F7 and CCL27) and skin2 triplex (LOR, KRT9 and CDSN) in conjunction with a housekeeping gene, HKG, triplex (B2M, UBC and UCE). The laboratories used different chemistries and instrumentation. All laboratories were able to successfully isolate and detect mRNA in contact traces (e.g., human skin, palm-, hand- and fingerprints, clothing, car interiors, computer accessories and electronic devices). The simultaneous extraction of RNA and DNA provides an opportunity for positive identification of the tissue source of origin by mRNA profiling as well as a simultaneous identification of the body fluid donor by STR profiling. The skin markers LCE1C and LOR and the housekeeping gene marker B2M were detected in the majority of contact traces. Detection of the other markers was inconsistent, possibly due to the low amounts and/or poor quality of the genetic material present in shed skin cells. The results of this and the previous collaborative RNA exercises support RNA profiling as a reliable body fluid/tissue identification method that can easily be combined with current STR typing technology.

RNA/DNA co-analysis from human menstrual blood and vaginal secretion stains: results of a fourth and fifth collaborative EDNAP exercise.

The European DNA Profiling Group (EDNAP) organized a fourth and fifth collaborative exercise on RNA/DNA co-analysis for body fluid identification and STR profiling. The task was to identify dried menstrual blood and vaginal secretion stains using specific RNA biomarkers, and additionally test 3 housekeeping genes for their suitability as reference genes. Six menstrual blood and six vaginal secretion stains, two dilution series (1/4-1/64 pieces of a menstrual blood/vaginal swab) and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 24 participating laboratories, using RNA extraction or RNA/DNA co-extraction methods. Two novel menstrual blood mRNA multiplexes were used: MMP triplex (MMP7, MMP10, MMP11) and MB triplex (MSX1, LEFTY2, SFRP4) in conjunction with a housekeeping gene triplex (B2M, UBC, UCE). Two novel mRNA multiplexes and a HBD1 singleplex were used for the identification of vaginal secretion: Vag triplex (MYOZ1, CYP2B7P1 and MUC4) and a Lactobacillus-specific Lacto triplex (Ljen, Lcris, Lgas). The laboratories used different chemistries and instrumentation and all were able to successfully isolate and detect mRNA in dried stains. The simultaneous extraction of RNA and DNA allowed for positive identification of the tissue/fluid source of origin by mRNA profiling as well as a simultaneous identification of the body fluid donor by STR profiling, also from old and compromised casework samples. The results of this and the previous collaborative RNA exercises support RNA profiling as a reliable body fluid identification method that can easily be combined with current STR typing technology.

RNA/DNA co-analysis from human saliva and semen stains--results of a third collaborative EDNAP exercise.

A third collaborative exercise on RNA/DNA co-analysis for body fluid identification and STR profiling was organized by the European DNA Profiling Group (EDNAP). Twenty saliva and semen stains, four dilution series (10-0.01 µl saliva, 5-0.01 µl semen) and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 20 participating laboratories using an RNA extraction or RNA/DNA co-extraction method. Two novel mRNA multiplexes were used: a saliva triplex (HTN3, STATH and MUC7) and a semen pentaplex (PRM1, PRM2, PSA, SEMG1 and TGM4). The laboratories used different chemistries and instrumentation and a majority (16/20) were able to successfully isolate and detect mRNA in dried stains. The simultaneous extraction of RNA and DNA from individual stains not only permitted a confirmation of the presence of saliva/semen (i.e. tissue/fluid source of origin), but allowed an STR profile of the stain donor to be obtained as well. The method proved to be reproducible and sensitive, with as little as 0.05 µl saliva or semen, using different analysis strategies. Additionally, we demonstrated the ability to positively identify the presence of saliva and semen, as well as obtain high quality DNA profiles, from old and compromised casework samples. The results of this collaborative exercise involving an RNA/DNA co-extraction strategy support the potential use of an mRNA based system for the identification of saliva and semen in forensic casework that is compatible with current DNA analysis methodologies.

Quantification of aortic valve area and left ventricular muscle mass in healthy subjects and patients with symptomatic aortic valve stenosis by MRI.

MRI allows visualization and planimetry of the aortic valve orifice and accurate determination of left ventricular muscle mass, which are important parameters in aortic stenosis. In contrast to invasive methods, MRI planimetry of the aortic valve area (AVA) is flow independent. AVA is usually indexed to body surface area. Left ventricular muscle mass is dependent on weight and height in healthy individuals. We studied AVA, left ventricular muscle mass (LMM) and ejection fraction (EF) in 100 healthy individuals and in patients with symptomatic aortic valve stenosis (AS). All were examined by MRI (1.5 Tesla Siemens Sonate) and the AVA was visualized in segmented 2D flash sequences and planimetry of the performed AVA was manually. The aortic valve area in healthy individuals was 3.9+/-0.7 cm(2), and the LMM was 99+/-27 g. In a correlation analysis, the strongest correlation of AVA was to height (r=0.75, p<0.001) and for LMM to weight (r=0.64, p<0.001). In a multiple regression analysis, the expected AVA for healthy subjects can be predicted using body height: AVA=-2.64+0.04 x(height in cm) -0.47 x w (w=0 for man, w=1 for female).In patients with aortic valve stenosis, AVA was 1.0+/-0.35 cm(2), in correlation to cath lab r=0.72, and LMM was 172+/-56 g. We compared the AS patients results with the data of the healthy subjects, where the reduction of the AVA was 28+/-10% of the expected normal value, while LMM was 42% higher in patients with AS. There was no correlation to height, weight or BSA in patients with AS. With cardiac MRI, planimetry of AVA for normal subjects and patients with AS offered a simple, fast and non-invasive method to quantify AVA. In addition LMM and EF could be determined. The strong correlation between height and AVA documented in normal subjects offered the opportunity to integrate this relation between expected valve area and definitive orifice in determining the disease of the aortic valve for the individual patient. With diagnostic MRI in patients with AS, invasive measurements of the systolic transvalvular gradient does not seem to be necessary.

[Selective increase of ventricular pacing threshold with complete exit block following DC cardioversion in a patient with a dual chamber pacemaker]. / Selektiver ventrikulärer Reizschwellenanstieg mit komplettem Exitblock nach elektiver externer Elektrokardioversion bei implantiertem Zweikammerschrittmacher.

We report the case of a 78-year old patient with a dual chamber pacemaker, who was admitted for cardioversion of atrial tachycardia. Transthoracic DC shock of 160 J was followed by transient loss of ventricular capture with complete exit-block and severe nodal bradycardia. Subsequent analysis of stimulation thresholds revealed a marked rise in the ventricular threshold only, whereas atrial threshold was unchanged. The selective dysfunction of ventricular capture is most likely caused by current-induced tissue damage at the electrode-endomyocardial interface by preferential shunting of high electrical energy into the ventricular lead as compared to the atrial lead. High output pacing prior to elective DC cardioversion is recommended to ensure consistent capture, particularly in pacemaker-dependent patients, and careful evaluation of pacemaker function after shock delivery should performed.

Nitric oxide attenuates the release of endothelium-derived hyperpolarizing factor.

BACKGROUND: The contribution of the endothelium-derived hyperpolarizing factor (EDHF), proposed to be a cytochrome P450-derived metabolite of arachidonic acid, to endothelium-dependent dilatation under physiological conditions has yet to be established, because its effect can be detected only after inhibition of NO synthase and cyclooxygenase. The possibility that NO exerts a feedback inhibition on EDHF formation was studied in isolated perfused arterial segments. METHODS AND RESULTS: Under combined blockade of NO synthase and cyclooxygenase, the EDHF-mediated vasodilatation elicited by receptor-dependent agonists in rabbit carotid and porcine coronary arteries was significantly attenuated by the NO donors C87-3786 and CAS 1609. The endothelium-independent dilatation elicited by isoproterenol was not altered by either NO donor. In NG-nitro-L-arginine-treated carotid artery segments, C87-3786 significantly attenuated the acetylcholine-induced increase in 6-keto-prostaglandin F1 alpha release, which was taken as an index of arachidonic acid liberation. In parallel experiments using cultured human endothelial cells, C87-3786 attenuated the Ca2+ response to bradykinin. The release of EDHF from a luminally perfused porcine coronary artery was detected by recording the membrane potential of downstream-situated cultured rat aortic smooth muscle cells. The NO donor C87-3786 had no effect on the hyperpolarization elicited by preformed EDHF but markedly inhibited its release from bradykinin-stimulated donor segments. CONCLUSIONS: These findings indicate that under physiological conditions, the production of EDHF is damped by NO. Therefore, it follows that when NO synthesis is impaired, alleviation of this intrinsic inhibition may, at least in part, maintain endothelial vasodilator function.

Lean body mass in twins.

A study of 49 pairs of monozygous (MZ) twins and 38 pairs of same-sexed dizygous (DZ) twins showed that lean body mass (LBM), as determined by potassium 40 counting, is under genetic influence. Intrapair variances for LBM are much smaller than those for body fat, which suggests that LBM has a higher degree of heritability. There is a correlation between the magnitude of intrapair LBM differences and intrapair weight differences for both sets of twins, showing that environment is also an important influence. The effect of weight variation on LBM variation is greater for thin people than for those with appreciable burdens of body fat, an observation previously made on individuals who undergo a nutrition-induced weight change.

Colonization of burns and the duration of hospital stay of severely burned patients.

In a retrospective study the influence of several factors on the length of hospital stay of severely burned patients (at least 24% total body surface area) has been investigated. The influence of these factors was studied by means of the Cox model survival analysis with time-varying covariates. Seventy-one patients were included in this study. The mean age was 32 years (range 1-82 years), the mean total body surface area burned 40% (range 24-80%) and the mean full-thickness area burned 32% (range 10-70%). The length of hospital stay was positively correlated with the extent of the burned area and with the age of the patient. Wound colonization with Enterobacteriaceae or with a combination of Pseudomonas spp. and Staphylococcus aureus was also associated with a prolonged stay in hospital.

Selective intestinal decontamination for prevention of wound colonization in severely burned patients: a retrospective analysis.

In this study the effect of selective intestinal decontamination of the digestive tract (SDD) on wound colonization was investigated. Ninety-one patients with at least 25 per cent total burned surface area (TBSA) were included in this study. All patients received oral polymyxin. In 63 patients oral co-trimoxazole and amphotericin B were added to the regimen. The addition of co-trimoxazole decreased the incidence of Enterobacteriaceae wound colonization from 71 per cent to 11 per cent (P less than 0.005). Colonization with Proteus was eliminated in patients treated with co-trimoxazole, compared with an incidence of 36 per cent in the group treated with polymyxin alone (P less than 0.001). The addition of amphotericin B decreased yeast colonization of the burn wound from 39 per cent to 10 per cent (P less than 0.005). A close relation was observed between burn wound colonization and colonization of the gastrointestinal tract. No resistant bacterial strains emerged during the period of study. These results suggest that SDD is an effective method for prevention of wound colonization. Further controlled studies are needed to establish the role of SDD in preventing burn wound colonization and wound sepsis.

Successful homografting in an elderly patient with extensive burns using his identical twin brother as skin donor.

A 60-year-old patient with 50 per cent body surface area (BSA) flame burns (of which 46 per cent were full skin thickness loss) was treated successfully with skin grafts taken from his identical twin brother to cover 20 per cent of the wounds. On theoretical grounds, it can be assumed that the period in hospital was reduced by 28 days. Blood for tissue and blood group typing should be taken at an early stage, before the patient is given a blood transfusion.

Effect of iron limitation on protein composition and ultrastructure of Haemophilus ducreyi.

Protein profiles of whole cells of Haemophilus ducreyi grown in the presence or absence of the iron chelator desferal, were compared by polyacrylamide gel electrophoresis. Each of four strains produced novel proteins in the range 43-160 kDa when cultured under conditions of reduced iron availability. At some sub-inhibitory concentrations, desferal produced enhanced growth, possibly due to it functioning as an exogenous siderophore. Organisms grown under conditions of reduced iron availability ultrastructurally showed also large periplasmic spaces between cytoplasm and outer membrane.

[Comparative anatomic studies of the tongue of Pan troglodytes (Blumenbach, 1799) and other primates. I. The chimpanzee tongue]. / Vergleichend-anatomische Untersuchungen an den Zungen von Pan troglodytes (Blumenbach, 1799) und weiteren Primates. Teil I: Befunde an der Schimpansenzunge.

This study was carried out on a comprehensive collection of tongues of Pan troglodytes. Material of tongues of Tupaia glis, Microcebus murinus, Chirogaleus major, Lemur catta, Galago crassicaudatus, Cebus apella, and Macaca mulatta were used for comparison of the different stages of the development of the papilla foliata. The tongue of Pan exhibits longitudinal growth correlated with the growth of length of the jaws and individual age. A typical foramen caecum as it occurs in man was gross anatomically not recognizable. Seldom a small circular depression appeared in the midline close to the papilla circumvallata centralis which could be the remnant of the foramen caecum. There is a remarkable accumulation of papillae filiformes and fungiformes at the apex linguae; the same seems to be true for other genera of primates. It is self-explanatory that the accumulation of papillae makes the tip of the tongue a particularly sensitive area; this has been found to be true even in man. One specimen of the prenatal age just before term was studied. It exhibited remarkably elongated, thread-like papillae, their function is unknown as yet. They are no longer present in a newborn specimen of 4 d of age. In the aboral part of the dorsum linguae, a special kind of papillae occur; they are flat lobes which carry moderately elongated, tiny processes at their free margin. The function of these papillae is unknown; taste buds are absent.

[Comparative anatomy of the tongue of Pan troglodytes (Blumenbach, 1799) and other primates. II. Findings in other primates]. / Vergleichend-anatomische Untersuchungen an den Zungen von Pan troglodytes (Blumenbach, 1799) und weiteren Primates. Teil II: Befunde an anderen Primaten.

This 2nd part of our studies shows that the papilla foliata is fully developed in Pan, Cebus, and Macaca; in Prosimians the papilla foliata is well developed in Lemur and Chirogaleus. In Galago crassicaudatus, this papilla is missing. Among 3 individuals of Microcebus, the papilla foliata was differently developed: in 2 cases, the tongue exhibited only 2 on both sides and a very low folia. Taste buds were found in the epithelium of only one side of each folium. In the 3rd case, the folia of the papilla were developed only on one side of the tongue, whereas, on the other side, a typical papilla was missing. Instead of the papilla, the tongue of the same animal exhibited a hillock-like structure; it is a gustatory hillock which exhibited many taste buds. There were 3 gustatory hillocks in all of the specimens of Tupaia glis; they are situated on both sides of the tongue.

Attempted suicide by means of burns.

A study was made of the results of treatment of 57 patients who made 68 suicide attempts by means of burns in the period between 1979 and 1987. They formed 14 per cent of the total patient population older than 16 years of age. Attempted suicide was observed most frequently in patients between 20 and 39 years of age. The sex incidence was equal. Most patients were in contact with or had been in contact with mental health care institutions at the time of their attempt, half of the attempts took place in a psychiatric hospital. Twelve patients died, in nine the prognosis was considered to be so poor that in view of the seriousness of the burns or the accompanying injuries no treatment was initiated. Follow-up took place after an average 3.8 years (1-8.25 years). Information was obtained on all 45 surviving patients. Eight patients had died, six of natural causes and two had committed suicide. Eighteen patients were living at home, 13 were residents at a psychiatric hospital, three were living in a protective commune, two were being detained during Her Majesty's pleasure and one was in prison. It appeared to be difficult to incorporate these patients in a closed follow-up regimen.

Disseminated melanomatosis of the central nervous system and other organs: a case report.

This case represents an example of clinically occult, disseminated neoplastic disease with almost asymptomatic but severe involvement of the central nervous system. The only helpful diagnostic investigation appeared to be cerebrospinal fluid cytology. Extensive post mortem examination was inconclusive as to whether the widespread CNS involvement with melanoma was primary or secondary.

Selective intestinal decontamination of the digestive tract for infection prophylaxis in severely burned patients.

An oral prophylactic antibiotic regimen aiming at suppression of the gram-negative rods and yeasts of the bowel flora was utilised in 48 severely burned patients to prevent burn wound colonisation. Only 17% of the patients had an actual or potential infection. Only one Pseudomonas infection occurred. The effect of this selective gastro-intestinal decontamination is discussed.

Psychological and social problems in burn patients after discharge. A follow-up study.

Approximately 18 months after discharge we were able to screen 42 burned adult patients for psychological and social problems. Using independent semi-standardized interviews, validated psychological questionnaires and standardized self-reports of the patients, we obtained three separate evaluations. The interreliability between the three different 'judges' varied from poor to moderate. Research suggests that 40% of burned adult patients have psychological problems. The cycle of recovery is dynamic. Only 21% need psychological help 18 months after discharge, but half of these are in danger of becoming 'drop-outs' because they do not receive the necessary treatment. A follow-up consisting of psychological questionnaires six months post-discharge appears to be a good and efficient instrument in limiting the drop-out risk.

Management of fingernail deformities after thermal injury.

Fingernail deformities after thermal injury can be very disfiguring and troublesome. The fingernails may be discoloured, cleft or vertically deviated. From the pathophysiology of the nail growth in general, the specific features of the nail deformities after deep burn can be deduced, as can guidelines for the treatment. Over a two-year period, 153 patients with burned hands were treated in the Groningen Burn Centre. Seven of these patients developed abnormal quality and growth pattern of the fingernails. In two patients an effort was made to correct the deformity by incising the dorsal skin at the base of the nail fold, reflecting the nail fold and covering the acquired defect with a full thickness skin graft. Although the patients were satisfied with the results achieved, one year after the operation little or no improvement in the nail growth was found. A cautious policy should be followed in correcting these deformities, since dislocation of the eponychium together with damage to the nail matrix and the nail bed play a complex role in the process of deformation.

Higher density of mental capacities in the left cerebral hemisphere of man: a quantitative investigation in children with localized cerebral lesions.

51 children with localized cerebral lesions were investigated with the British Ability Scales, Wechsler test, Porteus Mazes, Conners Teacher Rating Scale, and computerized tomography. The extent of the lesions was quantitatively determined. The IQ was 10 points lower in the left hemispheric lesions, although the right hemispheric lesions were larger. When corrected for size of lesions, the IQ is 17 points higher (p less than 0.05) in children with right than with left hemispheric early lesions. The children with left hemispheric lesions had also significantly more behavioural problems (Conners Scale). Children with bilateral lesions had 15 points lower IQ than those with unilateral lesions, although from the slightly larger extent one could expect only 3 points difference. There was no influence of hemiparesis or of epilepsy of the IQ independent of the extent of lesion. There was more plasticity of the hemisphere than of the lobes as shown by Wisconsin Card Sorting data. The higher density of mental capacities in the left cerebral hemisphere of man is explained by more efficient programs, and more dense packing of functions due to more training of the left cerebral hemisphere.