(E)-5-(2-bromovinyl)-2'-desoxyuridine--a new nucleoside analog with selective inhibitory action against herpesviruses. Studies in cell culture and animal experiments

(E)-5-(2-Bromovinyl)-2'-deoxyuridine (1; BrVUdR) inhibits the replication of herpes simplex virus type 1 (HSV-1) and of varicella-zoster virus (VZV) in vitro at concentrations of 0.01 to 0.23 mumol/l, whereas herpes simplex virus type 2 (HSV-2) is influenced only at 5.5 to 27 mumol/l. In comparison to some classical and newly developed antiherpetics, i. e. 5-iodo-2'-desoxyuridine (2; idoxuridine, IDU), 9-beta-D-arabinofuranosyladenine (4; vidarabine Ara-A), 9-(2-hydroxyethoxymethyl) guanine (5; acyclovir, ACV) and 2'-fluoro-5-iodo-1-beta-D-aracytosine (6;FIAC) the following order of decreasing activity was found:1 greater than 6 greater than 5 greater than 2 greater than 4 (against HSV-1) and 6 greater than 2 greater than 5 greater than 1 greater than 4 (against HSV-2). The high selectivity of the antiviral effect of BrVUdR towards HSV-1 and TZV is based on the fact, that proliferation of different mammalian cell lines is inhibited by 50% only at concentrations as high as 90 to 170 mumol/l, resulting in a therapeutical index of 1000 to 10,000. Successful treatment of an HSV-1 encephalitis in mice as well as an HSV-1 keratitis of rabbits confirmed the efficiency of 1 in experimental animal infections. No toxic side effects in both local and systemic applications were observed. Promising data from cell culture and animal experiments recommend 1 as a potential candidate for the local and systemic treatment of HSV-1 and VZV infections in man.     

Studies on the Influence of a Mutation of MASP-2 on the Binding to MBL and Ficolins

Chronic hepatitis B virus (HBV) infection affects about 200–400 million people worldwide and represents one of the leading causes for liver cirrhosis and hepatocellular carcinoma. Control over the HBV infection is achieved mainly by vaccination with Hepatitis B surface antigen (HBsAg). HBsAg contains N -linked glycosylation side and is recognized by both MBL-A and MBL-C in a Ca-dependent manner. HbsAg–MBL complexes activate complement and may thus affect humoural immunity. To investigate the role of MBL in humoural responses to HBsAg, we immununized mice that lack both MBL-A and MBL-C proteins with soluble HBsAg. It has been shown that deficiencies in other complement components like C1q, C4 and C3 result in decreased antibody responses. However, MBL double KO animals mounted dramatically increased humoural responses. After priming, MBL double KOs mounted HbsAg-specific IgM responses, which were threefold higher than WT controls. After boosting the HBsAg, total IgG was 10-fold higher in MBL KO than in WT control animals. Similar to the response to HbsAg, other glycosylated soluble antigens (e.g. invertase) induced better humoural responses in MBL double KO animals, suggesting that MBL plays an important role in a negative feedback regulation of adaptive immunity. Reconstitution experiments with rMBL partially rescued the KO phenotype. We propose that the clearance of glycoprotein antigens in MBL KO is handled differently from the WT, resulting in better stimulation of humoural responses. Alternatively, glycoprotein-Ag-MBL-rich complexes inhibit B-cell responsiveness via putative MBL receptors.     

Efficient high-frequency body coil for high-field MRI.

The use of body coils is favored for homogeneous excitation, and such coils are often paired with surface coils or arrays for sensitive reception in many MRI applications. While the body coil's physical size and resultant electrical length make this circuit difficult to design for any field strength, recent efforts to build efficient body coils for applications at 3T and above have been especially challenging. To meet this challenge, we developed an efficient new transverse electromagnetic (TEM) body coil and demonstrated its use in human studies at field strengths up to 4 T. Head, body, and breast images were acquired within peak power constraints of <8 kW. Bench studies indicate that these body coils are feasible to 8 T. RF shimming was used to remove a high-field-related cardiac imaging artifact in these preliminary studies. P41RR13230     

Amelioration of hypertrophic cardiomyopathy using nonsurgical septal ablation in a cirrhotic patient prior to liver transplantation.

A 53-year-old male with hepatitis C cirrhosis, who had been refused liver transplantation because of hypertrophic cardiomyopathy (HC), underwent nonsurgical septal ablation using alcohol with resolution of his ventricular outflow obstruction. This patient was able to subsequently undergo a successful deceased donor liver transplantation. This is the first reported case of alcohol induced septal ablation being performed in a cirrhotic patient with HC. Such nonsurgical procedures may be attractive in cirrhotic patients who are refused access to liver transplantation because of high surgical risk.     

Outcome in emotionally related living kidney donor transplantation

Background. The growing shortage of cadaver kidneys, the limited possibilities to expand the living related donor pool and the good results obtained in our centre with poorly matched cadaver kidneys, led us in 1991 to begin accepting highly motivated, unrelated, living kidney donors who had a strong emotional bond with the recipients. Methods. Between 1 January 1991 and 1 January 1996, 46 potential living kidney donors and their emotionally related recipients were evaluated. Twenty-three cases were accepted for renal transplantation after thorough somatic and psychological evaluation. The mean post-transplant follow-up until 1 April 1996 was of 28±3 months. Compatible blood groups and a negative cross-match were mandatory, but no minimal HLA matching was required. Results. There was a 50% drop-out rate following the initial screening. The main reasons for not performing transplantation were immunological contraindications in 39% of the cases, somatic in 30.5%, psychological in 26% and socioeconomic in 4.5%. In the accepted group of recipients, 48% (11/23) received transplants without chronic dialysis. Donor survival was 91%; two deaths unrelated to nephrectomy occurred 1 year after donation. The 2-year actuarial recipient and graft survivals were 100% and 91% respectively, compared to 99% (recipients) and 93% (grafts) in the non-HLA-identical living related kidney transplant group, and to 93% (recipients) and 83% (grafts) in the cadaver kidney transplant group. Recipient rehabilitation was completed after 4±1 months. Emotionally related donors returned to work 5±2 weeks after nephrectomy, and no donor regretted his decision, even in the case of failure. Conclusions. Kidney transplantation from emotionally related living donors represents a valuable option, allowing more patients with end-stage renal disease to avoid chronic dialysis. Recipient and graft outcome were superior to cadaver kidney transplantation. Motivated and emotionally related donors should be allowed to donate one of their kidneys provided that they are carefully selected and thoroughly informed.     

The incidence, timing, and management of biliary tract complications after orthotopic liver transplantation.

OBJECTIVE: This study analyzed the incidence and timing of biliary tract complications after orthotopic liver transplantation (OLTx) in 1792 consecutive patients. These results were then compared with those of previously reported series. Finally, recommendations were made on appropriate management strategies. SUMMARY BACKGROUND DATA: Technical complications after OLTx have a significant impact on patient and graft survival. One of the principal technical advances has been the standardization of techniques for biliary reconstruction. Nonetheless, biliary complications still occur. A 1983 report from the University of Pittsburgh reported biliary complications in 19% of all transplants, and an update in 1987 reported biliary complications in 13.2% of transplants. METHODS: The medical records of all patients who underwent liver transplantation and were hospitalized between January 1, 1988 and July 31, 1991 were reviewed. The case material consisted of the medical records of 217 patients treated for 245 biliary complications. RESULTS: Primary biliary continuity was established by either choledochocholedochostomy over a T-tube (C-C, n = 129) or a Roux-en-Y choledochojejunostomy with an internal stent (C-RY, n = 85). The overall incidence for biliary complication in this large series was 11.5%. Strictures (n = 93) and bile leak (n = 58) were the most common complications (69.6%). Most biliary complications (n = 143, 66%) occurred within the first 3 months after surgery. In general, leaks occurred early, and strictures developed later. Bile leaks were equally frequent in both C-C and C-RY (27.1% and 25.9%, respectively); strictures were more common after a C-RY type of reconstruction (36.4% and 52.9%, respectively). Twenty-one patients died, an incidence of 9.6%. Fifteen of the 21 biliary-related deaths were among patients treated for rejection before the recognition of biliary tract pathologic findings. CONCLUSIONS: Progress has been made on improving the results of biliary reconstruction after OLTx. Nonetheless, patients continue to experience biliary complications after OLTx, and these complications cause considerable loss of grafts and life. If significant additional improvement in patient and graft survival are to be obtained, the technical performance of OLTx must continue to improve.     

Recombinant human stem cell factor stimulates growth of a human glioblastoma cell line expressing c-kit protooncogene.

The growth of a panel of eight different human glioblastoma cell lines was examined in a human tumor cloning assay in agar, a tritiated thymidine uptake assay, and by counting cell numbers, in cultures performed in the absence or presence of increasing concentrations (1 to 100 ng/ml) of recombinant human stem cell factor (SCF). Growth of 7 of 8 cell lines was not significantly and reproducibly affected by recombinant human SCF. However, growth of the CRL 1620 cell line could be stimulated up to 5-fold by the cytokine. In contrast to the other cell lines investigated, CRL 1620 expressed the c-kit protooncogene assessed on the mRNA and protein level. Furthermore, SCF-induced proliferation of CRL 1620 cells was sensitive to the tyrosine kinase inhibitor erbstatin. Our data suggest that SCF can be operative in growth modulation of malignant cells outside the hematopoietic system, and this finding should be further studied for its possible clinical implications.     

DIALLELIC POLYMORPHISM MAY EXPLAIN VARIATIONS OF THE BLOOD CONCENTRATION OF MANNAN-BINDING PROTEIN IN ESKIMOS,BUT NOT IN BLACK AFRICANS

Mannan-binding protein (MBP) is a lectin which, upon binding to certain carbohydrates, activates the classical pathway of complement without the involvement of antibody or C1q. Deficiency of the MBP is associated with an opsonic defect and recurrent infections during early life. An amino acid substitution in the exon 1 at codon 54 in the MBP gene (GGC [glycine] to GAC [aspartic acid]) has been shown to be closely associated with low MBP concentration in Caucasoids. The gene frequency of the mutant allele in this population has been estimated at 0.13. In the study described here, we investigated the association between the mutant allele and MBP protein concentration in Eskimos from East-Greenland and black Africans from the Baringo District in Kenya. The frequency of the GAC allele was identical in Eskimos and Caucasoids (0.13). No overlap with regard to MBP concentration between the genotypes was found in the Eskimos. In contrast, the Africans revealed a low frequency of the GAC allele (0.009). However, the median MBP protein concentration was approximately 5 times lower among the Africans than the Eskimos. In 12.6% of the Africans and in 2.5% of the Eskimos, MBP was undetectable. Thus, MBP deficiency is the most frequent immunodeficiency so far described. The high prevalence of MBP deficiency among healthy individuals indicates that MBP deficiency also confers some selective advantages. We advance the hypothesis that MBP deficiency is maintained in populations because MBP deficiency decreases the infectivity of some intracellular micro-organisms which are dependent on opsonization.     

Validation in the sheep of an ultrasound velocity dilution technique for haemodialysis graft flow

BACKGROUND: A simple, rapid, inexpensive method for measuring the flow in a patient's vascular access would permit routine monitoring during haemodialysis, and hence provide information of access graft deterioration sufficiently early to increase the success of minimally invasive remedial procedures. This paper reports the validation of such a method in animals. METHODS: A PTFE graft was implanted in sheep between the carotid artery and the jugular vein. While the sheep was under general anaesthesia and on an haemodialysis circuit, ultrasound velocity in its blood was perturbed by the injection of a 5-10 ml bolus of isotonic NaCl. The pump tubing flow was measured by a transit-time blood flow meter. This flow was combined with the areas of perturbation generated by the injection before and after mixing in the access flow to estimate graft flow. The calculated graft flow was compared to flow measured directly by a transit-time probe on the same carotid artery. RESULTS: Over a 10-fold range, 120-1260 ml/min, graft flow measured by ultrasound velocity dilution agreed well with graft flow measured directly with a scatter of 76 ml/min about the regression line. CONCLUSION: Ultrasound velocity dilution provides a method for measuring flow in the graft accurate enough for clinical evaluation of patients on dialysis.      

An Assay for Mannan-Binding Lectin-Associated Serine Protease 3, MASP-3

In analogy to DNA microarrays, protein microarrays offer a new distinct possibility to perform sensitive high-throughput global proteome analysis. However, the development of the protein microarray technology will place high demands upon the design of both probes and solid supports. The analysis of thousands of heterogeneous proteins on a single microarray requires the use of uniform probes, such as antibodies, directly designed for protein microarray applications. We have recently generated a human recombinant single-chain Fv antibody library, genetically constructed around one framework, the nCoDeR-library, containing 2 × 1010 clones. Single framework antibody fragments (sinFabs) selected from this library were successfully applied as probes for microarrays providing sensitive detection in the 600 attomol (mass spectrometry) and the 300 zeptomole range (fluorescence). However, the choice of framework is critical. We have shown that the selected nCoDeR framework displayed excellent functional on-chip stability and arrayed dehydrated probes retained their activity for several months. Furthermore, we have addressed the issues of biocompatibility of the solid support and immobilization strategies for our microarray setup. An in-house-designed substrate, macroporous silicon coated with nitrocellulose (MAP3-NC7), displayed properties equal to, or better than, those of five commercially available supports used as reference surfaces. We have also evaluated different coupling strategies, such as adsorption, covalent coupling, diffusion and affinity coupling. Using a novel affinity tag, the double-(his)6-tag, we increased the binding efficiency of sinFab-molecules to Ni2⁺-coated solid supports, thereby allowing nonpurified probes to be directly applied.