Rotavirus capsid VP6 tubular and spherical nanostructures act as local adjuvants when co-delivered with norovirus VLPs.

A subunit protein vaccine candidate based on norovirus (NoV) virus-like particles (VLPs) and rotavirus (RV) VP6 protein against acute childhood gastroenteritis has been proposed recently. RV VP6 forms different oligomeric nanostructures, including tubes and spheres when expressed in vitro, which are highly immunogenic in different animal models. We have shown recently that recombinant VP6 nanotubes have an adjuvant effect on immunogenicity of NoV VLPs in mice. In this study, we investigated if the adjuvant effect is dependent upon a VP6 dose or different VP6 structural assemblies. In addition, local and systemic adjuvant effects as well as requirements for antigen co-delivery and co-localization were studied. The magnitude and functionality of NoV GII.4-specific antibodies and T cell responses were tested in mice immunized with GII.4 VLPs alone or different combinations of VLPs and VP6. A VP6 dose-dependent adjuvant effect on GII.4-specific antibody responses was observed. The adjuvant effect was found to be strictly dependent upon co-administration of NoV GII.4 VLPs and VP6 at the same anatomic site and at the same time. However, the adjuvant effect was not dependent on the types of oligomers used, as both nanotubes and nanospheres exerted adjuvant effect on GII.4-specific antibody generation and, for the first time, T cell immunity. These findings elucidate the mechanisms of VP6 adjuvant effect in vivo and support its use as an adjuvant in a combination NoV and RV vaccine.

Enantioselective and nonlinear intestinal absorption of eflornithine in the rat.

This study aimed to investigate if the absorption of the human African trypanosomiasis agent eflornithine was stereospecific and dose dependent after oral administration. Male Sprague-Dawley rats were administered single doses of racemic eflornithine hydrochloride as an oral solution (750, 1,500, 2,000, or 3,000 mg/kg of body weight) or intravenously (375 or 1,000 mg/kg of body weight). Sparse blood samples were obtained for determination of eflornithine enantiomers by liquid chromatography with evaporative light-scattering detection (lower limit of quantification [LLOQ], 83 microM for 300 microl plasma). The full plasma concentration-time profile of racemic eflornithine following frequent sampling was determined for another group of rats, using a high-performance liquid chromatography-UV method (LLOQ, 5 microM for 50 microl plasma). Pharmacokinetic data were analyzed in NONMEM for the combined racemic and enantiomeric concentrations. Upon intravenous administration, the plasma concentration-time profile of eflornithine was biphasic, with marginal differences in enantiomer kinetics (mean clearances of 14.5 and 12.6 ml/min/kg for L- and D-eflornithine, respectively). The complex absorption kinetics were modeled with a number of transit compartments to account for delayed absorption, transferring the drug into an absorption compartment from which the rate of influx was saturable. The mean bioavailabilities for L- and D-eflornithine were 41% and 62%, respectively, in the dose range of 750 to 2,000 mg/kg of body weight, with suggested increases to 47% and 83%, respectively, after a dose of 3,000 mg/kg of body weight. Eflornithine exhibited enantioselective absorption, with the more potent L-isomer being less favored, a finding which may help to explain why clinical attempts to develop an oral treatment have hitherto failed. The mechanistic explanation for the stereoselective absorption remains unclear.

Importance of pre-analytical factors contributing to measurement uncertainty, when determining sulfadoxine and sulfamethoxazole from capillary blood dried on sampling paper.

A bioanalytical method is developed and validated for determination of sulfadoxine (SD) and sulfamethoxazole (SM) in 100 microL capillary blood dried on sampling paper (Whatman 31ET Chr). SD and SM are extracted with 2000 microL perchloric acid and the liquid phase is loaded onto ENV+ solid-phase extraction columns. SD, SM, and the internal standard are separated on a Purospher STAR RP-18 liquid chromatography column (150 x 4.6 mm) with a mobile phase consisting of acetonitrile-sodium acetate buffer pH 5.2, I = 0.1 (33:67, v/v). Analytes are detected with UV at 256 nm. Lower limit of quantitation is 5 micromol/L, where precisions are 4.2% and 3.9% for SD and SM, respectively. Three brands of sampling papers have been compared with respect to absorption properties, extraction recoveries, and variations. Punching out dried blood spots (DBS) instead of cutting spots into strips prior to extraction has been evaluated by examining precision and accuracy of SD and SM determinations. Importance of uniformity of types of sampling paper, sampling volume and biological matrix, benefit of punching out discs from DBS, and impact on absorption properties of different brands of sampling papers are discussed. Avoiding pre-analytical errors whenever possible results in concentrations determined being more accurate and precise.

Determination of eflornithine enantiomers in plasma, by solid-phase extraction and liquid chromatography with evaporative light-scattering detection.

A bioanalytical method for determination of eflornithine (DFMO) in 1000 microL human plasma has been developed and validated. DFMO and the internal standard (IS) were analysed by liquid chromatography with evaporative light-scattering detection (ELSD). Separation was performed on a Chirobiotic TAG (250 mm x 4.6 mm) column with ethanol (99.5%):0.01 mol/L acetic acid-triethylamine buffer at the rate of 25:75% (v/v) with flow rate of 1.0 mL/min. For d-DFMO in plasma the inter-assay precision was 6.5% at 75 micromol/L, 6.6% at 375 micromol/L and 5.8% at 750 micromol/L. For l-DFMO in plasma the inter-assay precision was 10.4% at 75 micromol/L, 6.5% at 375 micromol/L and 5.0% at 750 micromol/L. The lower limit of quantification (LLOQ) was determined to 25 micromol/L where the precision was 4.3% and 5.7%, respectively.

Deoxythymidine pools of human skin and guinea pig organs.

The activity of antiviral nucleoside analogues like acyclovir is influenced by a number of cellular factors, one being the deoxythymidine (dThd) concentration. We therefore analysed the dThd concentration in human plasma and skin and in organs of guinea pig, the common experimental animal. High-performance liquid chromatography showed low amounts of dThd in human skin, 0.20-1.15 nmol/g, whereas guinea pig skin and spleen had 20-30 nmol/g and the concentration in guinea pig plasma was 10-times higher than in human plasma. These animals are therefore in this respect less suitable as accurate models for antiviral nucleoside activity in humans.

Methicillin-resistant Staphylococcus aureus in two burn units: clinical significance and epidemiological control.

Routine isolation adequately controlled MRSA strains in two burns units with a restrictive antibiotic policy. Ventilation control and more rigorous change of clothing offered no further advantage. No carriers among staff were found, but some suffered minor skin lesions that were the source of further MRSA spread. Spread of MRSA from the unit to other parts of the hospital was prevented by early identification of colonized patients and by restricting patient and staff movement.

Automated solid-phase extraction method for the determination of piperaquine in capillary blood applied onto sampling paper by liquid chromatography.

A bioanalytical method for the determination of piperaquine in 100 microL blood applied onto sampling paper, by solid-phase extraction and liquid chromatography, has been developed and validated. Blood spots were cut into small pieces prior to addition of 0.3M perchloric acid, acetonitrile and phosphate buffer containing an internal standard. The liquid phase was loaded onto a mixed phase cation-exchange (MPC) solid-phase extraction column. Piperaquine and the internal standard were analysed by liquid chromatography and separated on a Chromolith Performance (100 mm x 4.6 mm) column with acetonitrile:phosphate buffer pH 2.5, I = 0.1 (8:92, v/v) at the flow of 3.5 mL/min. The UV detection was performed at 345 nm. The intra-assay precision was 12.0% at 0.150 microM, 7.3% at 1.25 microM and 7.3% at 2.25 microM. The inter-assay precision was 1.8% at 0.150 microM, 5.2% at 1.25 microM and 2.8% at 2.25 microM. The lower limit of quantification (LLOQ) was determined to 0.050 microM where the precision was 14.7%.

Calcitonin gene-related peptide increases the blood flow of port-wine stains and improves continuous-wave dye laser treatment.

Ten patients with port-wine stains were treated with continuous-wave dye laser before and after treatment with the vasodilatory compound calcitonin gene-related peptide. The patients were given calcitonin gene-related peptide as an intravenous infusion or as a regional intravenous infusion. Six uniformly colored test sites, each 5 x 10 mm, located within a port-wine stain were selected for clinical evaluation. Laser treatment of these areas was performed on one occasion with three different energy densities: 15, 30, and 45 J/cm2. The port-wine stain circulation was monitored with a laser Doppler flowmeter before and after calcitonin gene-related peptide infusion and immediately after laser treatment. The cosmetic result was evaluated and judged as excellent, desirable, or undesirable. It was shown that calcitonin gene-related peptide is able to induce a long-lasting increase in blood flow in the vascular bed of port-wine stains and that the immediate postlaser laser Doppler flow values may serve as an indication of the outcome of laser therapy. The cosmetic result was often better in the group treated with calcitonin gene-related peptide compared with controls, and an excellent result also was achievable with the use of lower levels of energy.

A prospective and randomized study, "SVEA," comparing effects of three methods for delayed breast reconstruction on quality of life, patient-defined problem areas of life, and cosmetic result.

During the last 30 years, many methods for delayed breast reconstruction have been described. There is a lack of prospective randomized trials comparing reconstruction methods. The present study (SVEA), conducted 1995 to 1996, describes the impacts of three methods: the lateral thoracodorsal flap, the latissimus dorsi flap, and the pedicled transverse rectus abdominis muscle flap (TRAM), on important areas of life, patients' perception of cosmetic result, and quality of life. Questionnaires were completed before randomization and at 6 and 12 months postoperatively. The preoperative questionnaire concerned the impact of breast loss and expectations on reconstruction. Follow-up questionnaires dealt with satisfaction with cosmetic result and impact on important areas of life. A health-related quality-of-life questionnaire (SF-36) was completed at all points of assessment. A total of 75 of 87 randomized patients underwent breast reconstruction: 16 patients with the lateral thoracodorsal flap, 30 with the latissimus dorsi flap, and 29 with the TRAM flap. The majority were very satisfied with the cosmetic result. Most women reported improvements in important areas of life, and quality of life in terms of "social functioning" and "mental health" increased significantly after the reconstruction. The latissimus dorsi flap and TRAM flap scored significantly higher as compared with the lateral thoracodorsal flap for similarity with the contralateral breast and reduced problems in social situations. No differences between irradiated and nonirradiated patients were found. All methods were considered to produce good cosmetic results and improvements in patient-defined problem areas of life and quality of life. No negative effects were recorded. Thus, irrespective of method, breast reconstruction is a valuable tool for the mastectomized woman to cope with problems in everyday life.

Skin thickness in expanded human breast skin.

By means of an ultrasonic technique, skin thickness was measured in 13 women subjected to mammary reconstruction by tissue expansion. Measurements were performed the day before introduction of the expander, after the last inflation, the day before introduction of the permanent implant, and at 2 weeks and 6 months after this operation. Five patients had their expanders inflated every day (rapid expansion), and 8 patients, once a week (slow expansion). A decrease in skin thickness during the inflation period was recorded, and this decrease was permanent even 6 months after the second operation. After exchange of the expander for a permanent implant, there was a transient statistically significant increase in skin thickness. There was no statistical difference between the groups of rapid and slow expansion.

Ectatic blood vessels in port-wine stains lack innervation: possible role in pathogenesis.

The innervation pattern of port-wine stains was investigated using indirect immunohistochemistry with antibodies to protein gene product 9.5 (PGP 9.5), neuron-specific enolase (NSE), calcitonin gene-related peptide (CGRP), and neurofilament (NF). The pathologically dilated vessels in the middle and deep dermis were found to have defective innervation with only single or no nerve fibers in their vicinity, while other structures in the skin showed a normal density of fibers. NSE- and PGP-like immunoreactive (-LI) nerve fibers were observed innervating vessels with a normal morphology and other structures in the skin, such as sweat glands and hair follicles, as free nerve endings and in nerve bundles. The nerve bundles were often seen to pass the ectatic vessels without giving off any branches. CGRP-LI nerve fibers were detected running toward epidermis, whereas no fibers were found around the ectatic vessels. NF-LI fibers were seen innervating normal vessels in dermis, while in relation to the dilated vessels, no or only occasional fibers were observed. The lack of innervation may be of importance for the development of the disease as a result of decreased tonus of the vessels and/or a loss of neuronal trophic factors.

Alterations in skin properties during rapid and slow tissue expansion for breast reconstruction.

This study comprises 23 women who had had mastectomies because of breast cancer. They were randomly divided into two groups when they were admitted for breast reconstruction by tissue expansion. The first group was expanded rapidly, i.e., every day, and the other group was expanded slowly, i.e., every week. There were no other differences in the treatment between the two groups. Three months after completion of expansion, the expander was replaced by a permanent prosthesis. The follow-up time was up to 6 months after the second operation. Three different parameters--distensibility, elasticity, and hysteresis--were measured noninvasively on the breast skin and at a control site on several occasions throughout the treatment. During the treatment period there were no differences in skin properties between rapidly and slowly expanded patients. Of the three parameters, distensibility showed the most prominent changes: decreasing during the expansion period, increasing after the expander had been replaced by a permanent prosthesis, and decreasing during the following 6 months. Elasticity did not change significantly, except decreasing after insertion of the permanent prosthesis, and the hysteresis increased at the same time. These findings indicate that tissue expansion alters breast skin only to a small extent and that the mechanical resistance sometimes encountered during tissue expansion is due to deeper structures such as underlying muscles or capsule formation.

The inflammatory reaction in elective flap surgery.

The inflammatory response in three different flap procedures was investigated by measuring the preoperative and postoperative levels of C-reactive protein, leukocyte count, and body temperature. Patients scheduled for delayed breast reconstruction were operated on with the lateral thoracodorsal flap, the latissimus dorsi flap, or the pedicled TRAM flap. All patients received 2 gm of intravenous cloxacillin for antibiotic prophylaxis and 1 gm of paracetamol four times a day as basic treatment for postoperative pain. Within each treatment group, significant postoperative changes in C-reactive protein levels, leukocyte count, and body temperature were noted when compared with preoperative values. The highest C-reactive protein level (130 mg/ml) was found in the TRAM group on the third postoperative day. The kinetic pattern of C-reactive protein was similar for the latissimus dorsi flap and lateral thoracodorsal flap procedures, but the maximum C-reactive protein levels were significantly lower, 74 and 44 mg/ml respectively. Small (0.5 to 0.9 degrees C) but significant differences in body temperature were also noted on the second and third postoperative day. The TRAM flap group had the highest, the latissimus dorsi flap group intermediate, and the lateral thoracodorsal flap group the lowest value. The postoperative C-reactive protein levels seem to reflect the extent of the surgical trauma.

Effect of low power gallium arsenide laser on healing of venous ulcers.

The healing of venous ulcers of the leg with and without gallium arsenide laser treatment was studied in 42 patients randomly divided into two groups. One group received standard conservative treatment and gallium arsenide laser, and the other received the same standard treatment and placebo laser treatment. There were no differences in results between the two groups.

Hamartoma of the breast: surgical treatment and reconstruction. Case report.

Hamartoma of the breast is a rare and usually benign tumour with a variable growth pattern. Slowly increasing breast asymmetry is the most common clinical finding. Smaller hamartomas are often diagnosed on mammography. Simple enucleation and secondary expansion of the breast tissue are often sufficient to restore breast symmetry but sometimes, as in the two cases presented, reconstruction is necessary.

Continuous wave (CW) dye laser vs. CW argon laser treatment of port wine stain (PWS).

Pulsed dye laser treatment has shown specific vessel damage and thrombosis and good clinical bleaching of PWS at lower energy densities than continuous wave (CW) argon laser treatment. Based on a pilot study with CW dye laser treatment of PWS, we wanted to investigate whether CW dye laser would be more beneficial in PWS treatment than argon laser treatment. The hypothesis was tested in 20 randomized PWS patients, but no difference in treatment results was detected between the two treatment modalities.

Reactivity of the microcirculation in port wine stain hemangiomas evaluated by laser Doppler fluxmetry.

Twelve patients with port wine stains (both untreated and treated with argon laser) had their microcirculation monitored by laser Doppler flux at rest and after arterial occlusion lasting one minute. The blood cell flux was measured in normal, untreated and treated skin areas. The resting flux was significantly higher in untreated port wine stains than in the normal skin. The reactive hyperemia response was significantly impaired in the untreated skin, although a 60% increase in blood cell flux was achieved by the short arterial occlusion. In most of the recorded parameters the treated areas showed a less pathological reaction than the untreated ones. We conclude that peroperative vasodilation can be induced in port wine stains, and this may be used to improve the outcome of argon laser treatment.

The lateral thoracodorsal flap in breast reconstruction: a comparison between two plastic surgical centres.

In a retrospective study the results of breast reconstruction with 146 lateral thoracodorsal flaps operated on during the period 1991-94 at the Departments of Plastic Surgery at the Sahlgrenska University Hospital in Göteborg and the Stockholm Söder Hospital were evaluated. The median age of the 135 patients was 51 years (range 37-74). About two thirds of the patients were also operated on on the opposite side at the same time; and this was done more often in Stockholm, which partly explains the longer operating time in Stockholm. Perioperative bleeding (median 100 ml, range 25-400) was similar in both groups. Median postoperative bleeding, which was measured only in Stockholm, was 300 ml (range 30-1000). Seromas were seen only in Göteborg where postoperative drains were rarely used. The rate of early complications (infection and partial necrosis) were higher in Stockholm where more flaps were raised in irradiated tissue. We conclude that the results were similar in the two centres, and the thoracodorsal flap gives good results. The incidence of early morbidity could be reduced by excluding patients who had been irradiated, by inserting drains, and by giving antibiotic prophylaxis.

Clinical follow up of the lateral thoracodorsal flap in breast reconstruction: comparative evaluation from two plastic surgical centres.

A follow up study that compared the results from two plastic surgical centres comprised 95 patients after breast reconstruction with the lateral thoracodorsal (LTD) flap combined with either a smooth or textured saline-filled implant. It was conducted a median of 39 months (range 15-67) after the reconstruction. We investigated the size of the reconstructed breast and LTD flap, symmetry of the breasts, orientation of the mastectomy scar, and the rate of capsular contracture, which were similar in the two centres. However, according to the modified Baker classification and applanation tonometry reconstructions with smooth-surfaced implants and drainage of the implant pocket resulted in softer breasts (p = 0.03). The LTD flap technique in breast reconstruction was a good choice, particularly for women who required a minor breast reconstruction, and the technique can be recommended for suitable patients.

Argon laser treatment of port wine stain.

The authors achieved excellent results using argon laser treatment of PWS in 45% of 84 patients. Punch biopsies were taken preoperatively in all patients. Their classification according to fullness of erythrocytes of the vessels was found to be of relative value in predicting the treatment result. It is thus found to be advisable to administer a test treatment of a limited area in all patients.