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Dendritic cells (DCs) are currently being considered as adjuvants in immunotherapy. Depending on their source and culture conditions, they show different features and maturation states. Dendritic cells can be generated from monocytes and CD34+ haematopoietic stem cells, from both adult and cord blood. Here, we report the generation of mature DCs from enriched CD34+ cord blood (CB) cells using autologous cord blood plasma (ACBP) as a source of serum proteins and factors. In the presence of ACBP, CD34+ cells proliferated and differentiated resulting in a population of cells with a dendritic phenotype as assessed by morphology and flow cytometry analyses. The DC population obtained using ACBP showed higher levels of HLA class II molecules, co-stimulatory molecules including CD40, CD80 or CD86, and the dendritic cell marker CD83, compared with those generated in adult blood serum (ABS). Furthermore, the DCs generated in the presence of ACBP were more potent stimulatory cells in the mixed lymphocyte:dendritic cell reactions (MLDCR), compared to cells generated in ABS. Similar results were obtained using homologous cord blood plasma (HCBP). These results show that ACBP can support the generation of DCs from CD34+ progenitor cells when only GM-CSF and TNFalpha are used as differentiating cytokines.

Molecular cloning techniques and V gene phage display have revolutionised the production of human monoclonal antibodies. Antibodies of a defined specificity can be obtained by selecting phage display libraries on antigen in a process known as panning. We have applied these techniques to the isolation of three HLA-A2-specific single chain variable domain fragments (scFv) from a patient alloimmunised by blood transfusion. Analysis of specificity with cells of HLA genotyped donors revealed the following: i) in addition to the major reactivity with HLA-A2, cross-reactivity with the HLA-A28 epitope; and ii) inhibition of scFv binding to the antigen by the patients' antibodies. The heavy chain variable genes of all three were derived from the germline gene Cos-3, carry the hallmarks of somatic hypermutation, and are most likely derived from clonally related B cells. The light chain variable domains were encoded by DPK1 and DPK8 from the VkappaI family. These data show that phage display can be used to clone HLA-specific alloantibodies that recognise the native antigen from alloimmunised patients.

The World Marrow Donor Association has formulated guidelines for establishing the extent and quality of histocompatibility testing for unrelated donor registries, umbilical cord blood banks, and transplant centers involved in international exchange of hematopoietic stem cells for allogeneic transplantation. The ability to identify unrelated stem cell donors in one country for patients in another country requires cooperation and standardization in many areas. The adoption of guidelines for histocompatibility testing, such as those summarized in this report, will facilitate these opportunities and rapidly provide accurately typed donors for patients in need.

The London Cord Blood Bank was established with the aim of collecting, processing and storing 10000 unrelated stem cell donations for the significant number of children in the UK requiring transplantation, for whom a matched unrelated bone marrow donor cannot be found. Collection is performed at two hospitals by dedicated cord blood bank staff after delivery of the placenta. Mothers are interviewed regarding medical, ethnic and behavioural history by nurse counsellors and sign a detailed consent form. Donations are returned to the bank for processing. Volume reduction is undertaken by a simple, closed, semi-automated blood processing system, with excellent recovery of progenitor cells. Units are cryopreserved and stored in the vapour phase of liquid nitrogen. Blood samples from mothers and cord blood donations are tested for the UK mandatory red cell and microbiology markers for blood donors. Donations are typed for HLA-A, B and DR at medium resolution (antigen split) level using sequence-specific oligonucleotide probing and sequence-specific priming techniques. The selection of collection hospitals on the basis of ethnic mix has proven effective, with 41.5% of donations derived from non-European caucasoid donors. Bacterial contamination of collections has been dramatically reduced by implementation of improved umbilical cord decontamination protocols.

The World Marrow Donor Association has formulated guidelines for establishing the extent and quality of histocompatibility testing for unrelated donor registries, umbilical cord blood banks, and transplant centers involved in international exchange of hematopoietic stem cells for allogeneic transplantation. Registry and cord blood bank guidelines suggest that, at a minimum, initial HLA typing should be performed for three HLA loci, HLA-A, -B, and -DR, at low resolution/split antigen level. DNA-based testing methods should be utilized for HLA-DR typing. DNA-based testing for HLA-A and -B should replace serologic testing of new volunteer donors and cord blood units as robust protocols and reagents become available to the laboratories. Transplant center guidelines for typing of patient, family and to confirm the HLA types of potential unrelated donors should include, at the minimum, typing HLA-A, B, and -DR loci using primarily DNA-based testing methods at allele level resolution for DRB1 and low resolution/ split antigen level for HLA-A and -B. It is strongly recommended that the typing of a patient and the selected donor be performed using the same set of reagents, methodology, and interpretation criteria with fresh tissue samples to ensure HLA identity. Guidelines for laboratory accreditation, approaches to quality assurance and quality control for HLA testing, and suggestions for the format of the HLA database of donor types are also outlined.

The World Marrow Donor Association has formulated guidelines for establishing the extent and quality of histocompatibility testing for unrelated donor registries, umbilical cord blood banks, and transplant centers involved in international exchange of hematopoietic stem cells for allogeneic transplantation. Registry and cord blood bank guidelines suggest that, at a minimum, initial HLA typing should be performed for three HLA loci, HLA-A, -B, and -DR, at low resolution/split antigen level. DNA-based testing methods should be utilized for HLA-DR typing. DNA-based testing for HLA-A and -B should replace serologic testing of new volunteer donors and cord blood units as robust protocols and reagents become available to the laboratories. Transplant center guidelines for typing of patient, family and to confirm the HLA types of potential unrelated donors should include, at the minimum, typing HLA-A, B, and -DR loci using primarily DNA-based testing methods at allele level resolution for DRB1 and low resolution/split antigen level for HLA-A and -B. It is strongly recommended that the typing of a patient and the selected donor be performed using the same set of reagents, methodology, and interpretation criteria with fresh tissue samples to ensure HLA identity. Guidelines for laboratory accreditation, approaches to quality assurance and quality control for HLA testing, and suggestions for the format of the HLA database of donor types are also outlined.

Clinical evidence indicates that placental/umbilical cord blood (CB) is an alternative source of haematopoietic stem cells for bone marrow reconstitution. To establish a CB bank large panels of frozen, HLA-typed CB units need to be stored. Cryopreserved, unprocessed CB units require vast storage space. This study describes a method, using the Optipress II Automated Blood Component Extractor (Opti II) from Baxter Healthcare Corporation, to reduce the volume of the CB collection, preserving the quantity and quality of the progenitor cells, in a closed system. The CB collection was transferred to a triple bag system, centrifuged to produce a buffy coat layer and processed using a standard Opti II protocol to separate the whole blood into three components: plasma, buffy coat and buffy coat-depleted red cell concentrate. The buffy coat volume was standardised to 25 ml; mean reduced volume of 24.5 ml (s.d. 1.5 ml) with 53% red cell depletion. Good recovery of cells was observed: 92%, 98%, 96% and 106% recovery of nucleated, mononuclear, CD34+ and total colony-forming cells, respectively. Using this method for processing CB units reduces storage requirement by two-thirds but preserves the quantity and quality of the progenitor cells.

Advances in cannulation techniques and instruments have helped in difficult bile duct cannulation and thus stone extraction. For small common bile duct (CBD) stones, endoscopic papillary balloon dilatation has been proposed as an alternative to endoscopic papillotomy (EPT). The technique must undergo further evaluation before recommending its routine use. For most patients with bile duct stones, EPT remains the method of choice. Out of 8204 patients treated in three surgical endoscopy centers (Chile, Germany, and India), 86% to 91% of all CBD stones could be extracted subsequently after EPT using a Dormia basket; 4% to 7% required mechanical lithotripsy (ML) before removal and 3% to 10% of the patients needed other sophisticated techniques, such as electrohydraulic lithotripsy (EHL), laser-induced shock-wave lithotripsy (LISL), or extracorporeal shock-wave lithotripsy (ESWL). The local expertise and availability of equipment determines the choice of method used. In general, EHL or LISL is used for impacted CBD stones including stones in Mirizzi syndrome refractory to ML. ESWL is best suited for intrahepatic stones. Permanent stenting can be offered to poor risk patients instead of extensive procedures to clear the bile duct. Using currently available nonsurgical techniques, fewer than 1% of all patients with bile duct stones still require surgical intervention.

Umbilical cord blood (CB) transplantation is thought to be associated with a reduced risk of severe graft-versus-host-disease (GVHD) compared with bone marrow transplantation (BMT). The cytokine cascade is known to be important in the pathogenesis of GVHD; however, previous studies investigating the cytokine secretion pattern of CB cells have been contradictory because of variations in experimental techniques. In this study, the cytokine profile of cord and adult blood lymphocytes and lymphocyte subsets has been assessed at the single-cell level by flow cytometry, using CD4/CD8 and CD45RA/CD45RO markers. Cord and adult blood mononuclear cells were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin in the presence of monensin. After 4 to 24 hours of incubation, interleukin-2 (IL-2), IL-4, interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha (TNF-alpha) production was measured by three-color flow cytometry. The results show that cord blood lymphocytes (CBL) produce less IL-2, IL-4, IFN-gamma, and TNF-alpha than adult peripheral blood lymphocytes (ABL). Further subset analysis showed that in CBL the majority of cytokine producing cells were CD4(+)CD45RA+, whereas in ABL the cytokine-producing cells were both CD4(+)CD45RO+ and CD8(+)CD45RO+. These results suggest that the reduced incidence of GVHD in CB transplantation may partly due to the altered cytokine profile seen in CBL.

Cord blood (CB) contains a sufficient number of haemopoietic progenitor cells and can be used as an alternative to bone marrow transplantation. To date, a number of transplants using cord blood-derived stem cells from HLA-identical siblings and unrelated donors have been successfully performed. However, only approximately 30% of patients have an HLA-identical sibling while the rest are reliant on finding an HLA-compatible unrelated donor. This is particularly difficult for patients from ethnic minorities since the majority of unrelated bone marrow donor volunteers are from Caucasoid ancestry. The London Cord Blood Bank was established to provide an additional source of stem cells for all patients requiring bone marrow transplantation.

Interactions involving the T-cell receptor (TCR) and major histocompatibility complex (MHC) are fundamental to the generation of a specific immune response. The study of interpopulation differences in TCR genes may identify those genes which are subject to selection, and also provides useful information for future genetic studies in these populations. In this study we present analysis of five TCRAV polymorphisms, for V5S1, V6S1, V8S1, V17S1, and V21S1 loci in five human populations by single-strand conformational polymorphism (SSCP) analysis. Caucasian, Chinese, Gambian, AfroCaribbean, and South American Indians (Mapuches) showed marked interpopulation variation for both the silent (V5S1, V17S1, and V21S1) and coding (V6S1 and V8S1) polymorphisms. In general the alleles were conserved in the different populations, but new, additional variants were found for V5S1 and V17S1 in Gambians and Caucasians. V6S1 overall showed the highest nucleotide diversity, and V6S1 genotype distributions were skewed away from expected values in Chinese and Mapuches. Analysis of allelic associations showed a general lack of linkage disequilibrium between the loci, which was reflected by the absence of strong population-specific haplotypes.

An essential prerequisite for successful procurement of sufficient autologous peripheral blood progenitor cells (PBPC) for engraftment is the optimal timing of collection. A number of surrogate markers of peripheral blood progenitor cells were analysed to identify a single test which could predict the optimum time to harvest, providing at least 2 x 10(6) CD34+ cells/kg patient body weight. The study comprised 95 patients undergoing varied mobilisation regimens with chemotherapy and G-CSF for both solid tumours and haematological malignancies. One hundred and fifty-seven PBPC harvests were collected. Full blood counts (FBC) and CD34+ cell enumeration was performed on blood samples taken during the mobilisation period and immediately prior to leucapheresis (pre-harvest). All PBPC collections were assayed for colony-forming cells and CD34+ cells in addition to a FBC. The white cell count on the day of harvest showed only weak correlation with the total number of CD34+ cells in the collection (r = 0.30). In contrast, the absolute number of circulating CD34+ cells strongly correlated with the CD34+ cell and CFU-GM yield of the corresponding apheresis product. Provided the mobilisation sample contained > or =20 x 10(6) CD34+ cells/ml, 94% of single collections, performed the following day, contained > or =2 x 10(6) CD34+ cells/kg.

Background: The prevalence of chronic pancreatitis in Chile is not known and there is no local information about the surgical treatment of this disease., Aim: To review retrospectively the results of surgical treatment of chronic pancreatitis., Material and Methods: The charts of 17 patients (12 male), aged 7 to 65 years old, with chronic pancreatitis that were operated in three different Chilean regions, were reviewed., Results: Seven patients had previous endoscopic therapeutic procedures (papillotomy in 4 and stent placing in 3). Seven patients had been subjected to previous biliary surgical procedures. Indications for surgery were severe pain in 14 patients, the suspicion of a pancreatic carcinoma in 4 patients, an infected pseudocyst in one and massive bleeding of multiple pseudo-aneurysms in a pseudocyst in one patient. Twelve patients were subjected to decompressions and 5 to pancreatic resections. There was no operative mortality and one transient pancreatic fistula. After an average follow up of 22 months, pain improved in 94% of cases, pancreatic cancer was diagnosed in one patient and 79% of subjects gained weight. One patient became insulin dependent, one increased his insulin requirements and one had transient steatorrhea, since she could not afford pancreatic enzyme replacement therapy., Conclusions: The multidisciplinary approach of patients with chronic pancreatitis, with selective use of surgery, may greatly improve their quality of life.

Pregnancy can be considered a successful transplantation of allogeneic paternal tissue to the mother. Soluble HLA class I serum levels have been found to increase during solid organ rejection episodes and during graft-versus-host disease after bone marrow transplantation. We wished to determine whether significant changes in sHLA class I and beta 2-microglobulin light chain levels occurred during pregnancy, because these may reflect adaptive changes permitting the acceptance of the fetal graft. Serum samples were obtained from women at different stages of pregnancy and in the postpartum period. Cord blood samples and serum samples from nonpregnant female and male controls living in the same geographic area in Southern Chile were also studied. The levels of sHLA class I heterodimers were determined by an ELISA sandwich technique; beta 2-microglobulin levels were measured by MEIA IMX-Abbott. There was a significant elevation of sHLA class I levels in the first 2 trimesters of pregnancy, followed by a significant drop below normal levels at the end of pregnancy, with normalization in the post-partum period. beta 2-microglobulin levels did not change significantly during pregnancy and did not correlate with sHLA class I levels. In cord blood samples, sHLA class I levels were lower and beta 2-microglobulin levels higher than those of adult controls and of mothers at the time of delivery. The variations in sHLA class I levels during pregnancy may reflect or contribute to immunoregulatory events related to the acceptance of the fetal graft.

A previously transfused female patient, known to have a platelet defect, was transfused with platelets prior to surgery. After the 18th unit she felt unwell, developed fever, rigor, became nauseous, and vomited. Her blood pressure decreased from 140/90 to 80/50 mm Hg. Passive transfer of donor granulocytes or red cell antibodies were excluded as a cause. Therefore, a serum sample from the patient was investigated for the presence of antibodies to human leukocyte antigens (HLA) using a standard National Institutes of Health (NIH) lymphocytotoxicity test, but antibodies were not detected. However, an extended incubation cytotoxicity test demonstrated the presence of an anti-HLA-A2, and indirect immunofluorescence flow cytometry showed the presence of an IgG1 antibody reacting with 50 percent of cells in a random pool of lymphocytes. One week later, multispecific HLA antibodies were detectable by both NIH and extended incubation cytotoxicity tests. Flow cytometry showed a 16-fold increase in the amount of IgG antibodies and the appearance of an IgM component. Such clinically important HLA antibodies can be detected by extended incubation cytotoxicity and flow cytometric assays prior to becoming reactive in a standard NIH cytotoxicity technique.

1. SCA40 (0.1 nM-0.1 mM) produced concentration-dependent suppression of the spontaneous tone of human isolated bronchus (-log EC50 = 6.85 +/- 0.09; n = 10) and reached a maximal relaxation similar to that of theophylline (3 mM). The potency (-log EC50 values) of SCA40 compared to other relaxants was rolipram (7.44 +/- 0.12; n = 9) > SCA40 > or = levcromakalim (6.49 +/- 0.04; n = 6) > SKF94120 (5.87 +/- 0.10; n = 9). 2. When tested against the activity of the isoenzymes of cyclic nucleotide phosphodiesterase (PDE) isolated from human bronchus, SCA40 proved highly potent against PDE III (-log IC50 = 6.47 +/- 0.16; n = 4). It was markedly less potent against PDE IV (4.82 +/- 0.18; n = 4) and PDE V (4.32 +/- 0.11; n = 4). 3. Human polymorphonuclear leukocytes (PMNs) stimulated with N-formylmethionyl-leucyl-phenylalanine (FMLP) produced a concentration-dependent superoxide anion generation and elastase release. SCA40 (1 nM-10 microM) produced a concentration-related inhibition of FMLP (30 nM approximately EC50)-induced superoxide production (-log IC50 = 5.48 +/- 0.10; n = 6) and elastase release (-log IC50 = 5.50 +/- 0.26; n = 6). Rolipram was an effective inhibitor of superoxide generation and elastase release (-log IC50 values approximately 8) while SKF94120 and levcromakalim were scarcely effective. 4. FMLP (30 nM) and thimerosal (20 microM) induced leukotriene B4 production and elevation of intracellular calcium concentration in human PMNs. The production of leukotriene B4 was inhibited by SCA40 in a concentration-related manner (-log IC50 = 5.94 +/- 0.22; n = 6) but SCA40 was less effective against the elevation of intracellular calcium. Rolipram was an effective inhibitor of leukotriene B4 synthesis (-log IC50 approximately 7) and intracellular calcium elevation (-log IC50 approximately 6) while SKF94120 and levcromakalim were scarcely effective. 5. It is concluded that SCA40 is an effective inhibitor of the inherent tone of human isolated bronchus. The bronchodilatation produced by SCA40 appears mainly related to PDE inhibition since the potency of SCA40 as a relaxant of human isolated bronchus was found to be close to its potency as inhibitor of PDE III activity isolated from human bronchus. In addition, SCA40 exhibited inhibitory effects on human PMN function stimulated by FMLP. These effects may be related to the ability of SCA40 to inhibit PDE IV from human PMNs while the contribution of PDE V inhibition is uncertain. We found no evidence of a role for levcromakalim-sensitive plasmalemmal K+-channels in human PMNs.

Phosphodiesterase (PDE) isoenzyme type IV is the predominant cyclic AMP hydrolytic activity in polymorphonuclear leukocytes (PMNs). PDE IV inhibitors depress functional responses of PMNs but their influence on intracellular calcium concentration ([Ca2+]i) has not been extensively studied. The present study examined the effects of rolipram (a selective PDE IV inhibitor) on the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (fMLP)-induced changes of [Ca2+]i in fura-2 loaded human PMNs. Rolipram (1 nM-10 microM) did not alter basal [Ca2+]i values. fMLP (10 nM approximately EC50) produced a transient calcium response, i.e., a peak followed by decay to a residual value above baseline. Peak [Ca2+]i values after fMLP were not altered but a faster decay and a lower residual [Ca2+]i were observed in rolipram (0.1-10 microM)-treated cells. fMLP added after thimerosal (20 microM) produced a peak followed by a sustained oscillatory response. Rolipram (up to 10 microM) did not alter the peak but inhibited the sustained response (-log IC50 = 6.39 +/- 0.12). The inhibitory effects of rolipram may be due to alterations in the mobilization of Ca2+ produced by the increase in the cellular content of cyclic AMP. SKF94120 (a selective PDE III inhibitor) produced minor effects on the fMLP-induced calcium response. SCA40 (a mixed PDE III/IV/V inhibitor) produced similar effects but was less potent than rolipram. Reduction of the calcium response probably underlies the inhibition of PMN functions produced by PDE IV inhibitors.

It has been established that oligoclonal expansion is a common feature of the CD8+ T cell population, particularly within the CD8+ CD57+ lymphocyte subset. In addition, clonal malignancies involving CD8+ CD57+ T cells (large granulocytic lymphocytic leukemias) are often accompanied by rheumatoid arthritis, Felty's syndrome, or both. Therefore, to identify disease-related alterations in the CD8+ T cell repertoire, we have compared the patterns of oligoclonality in the CD8+ T cells of rheumatoid arthritis patients (n = 32) with those of age-matched controls (n = 25). By using a multiplex PCR assay for the CDR3 length of TCR beta-chains, we have found a striking increase in the frequency of CD8+ oligoclonality involving V beta 3 TCR: 50% of the rheumatoid arthritis patients had evidence of oligoclonality in this TCR family compared with 4% of controls (p < 0.0002). In addition, two unrelated RA patients had clonally dominant CD8+ T cell beta receptors that were identical in amino acid sequence, suggesting selection by a common Ag. An analysis of a subset of RA patients with mAbs specific for V beta 3 TCR revealed the presence of clonal expansion in a minority of patients usually, but not exclusively, involving the CD57+ subset. These data define a phenotype of the T cell repertoire that is strongly associated with rheumatoid arthritis; the mechanisms and genetic and environmental factors that explain this phenomenon remain to be defined.

In recent years the development of recombinant DNA and sequencing techniques has led to a greatly increased understanding of the genetic complexity, structure and function of the human major histocompatibility complex. This system may be subdivided into the "classical' HLA (human leukocyte antigen) class I and II transplantation antigens and novel HLA and non-HLA genes, involved in antigen processing and presentation to T cells. Parallel technological developments in HLA DNA typing in the clinical laboratory have resulted in a more precise awareness of the role of HLA matching for the classical HLA antigens in bone marrow and solid organ transplantation, while alternative strategies and techniques for donor selection are currently under evaluation. This review offers a current perspective on the genetics, structure and function of the HLA system, its relevance to clinical transplantation and future prospects for improvements in donor selection.

Previous reports have shown an increased frequency of certain HLA antigens in association with erythema multiforme, including HLA-B15(B62), HLA-B35, HLA-A33, HLA-DR53 and, more recently, HLA-DQB1*0301. A strong association with HLA-DQ3 has been documented in patients with recurrent erythema multiforme. We have performed HLA typing in 39 patients with recurrent erythema multiforme, of whom 33 were associated with herpes simplex virus infection. The results were compared with 309 controls. In the recurrent erythema multiforme patients there was a statistically significant increase in HLA-B62 and HLA-B35. An increase in HLA-DR53 was also found, although this did not reach statistical significance. There was no increase in HLA-A33. The presence of HLA-DQ3 in the study population approached that in the controls. Finally, the study population demonstrated a trend towards a reduction in the HLA antigens A1, B8 and DR3. The study confirms the previously reported associations with HLA-B62 (B15), HLA-B35 and HLA-DR53. We have been unable to confirm an association of HLA-A33 or HLA-DQ3 with erythema multiforme. The HLA antigens A1, B8, and DR3 are associated with autoimmune disease, reflecting an increased host response to tissue self antigens. Their absence in patients with recurrent erythema multiforme (REM) may be an indicator of a poor host response to an antigen, which in the case of REM is the herpes simplex virus.

Setting: Studies showing significantly higher concordance of tuberculosis among monozygotic twins than dizygotic twins have provided support for genetically determined susceptibility to tuberculosis., Objective: We wished to explore whether the development of delayed type hypersensitivity to tuberculin after newborn BCG immunization of twins suggested genetic regulation of the response to BCG in humans., Design: Our study population consisted of 17 monozygotic twin pairs, 18 dizygotic twin pairs, and 64 single infants 3-34 months of age from Santiago, Chile. All had a BCG scar and were tuberculin tested by one trained nurse., Results: The mean birth weight of both groups of twins was significantly lower than that of singletons and the percentage of individuals who failed to respond to tuberculin was approximately twice as high in twins as in singletons. After adjustment for birth weight and age by regression analysis, it was found that the distribution of tuberculin reactivity in both monozygotic and dizygotic twins was not significantly different from that of singletons. Both twin pair correlations is adjusted tuberculin reactivity were significantly greater than zero (P < 0.01) and led to a heritability estimate of 0.28. However, the monozygotic twin correlation was not significantly larger than the dizygotic twin correlation so that heritability is poorly estimated., Conclusion: These results are consistent with a genetic regulation of the response to newborn BCG immunization in humans by a mechanism capable of producing similar responses in identical and nonidentical twins alike.

Very early onset of chorea of Huntington is described in two sibs who have received the mutated gene from their affected father and grandfather. The consequences and the molecular mechanisms of germinal genomic imprinting and the amplification of the expanded CAG trinucleotide repeat are discussed.

An increased frequency of human leukocyte antigen (HLA)-DR1 was found in 49 non-immunosuppressed patients with non-melanoma skin cancer (NMSC), being highest in patients under the age of 60 with multiple squamous cell carcinomas (SCC). Of 266 patients receiving long-term immunosuppression following renal transplantation 46 (17%) were found to have NMSC. No increase in HLA-DR1 was found in renal transplant recipients (RTR) with non-melanoma skin cancer (RTR+C) when compared with matched renal transplant recipients without skin cancer (matched RTR-C), or when compared with healthy controls. There was an increased frequency of DQw2 in RTR+C, most pronounced in RTR with SCC (61.9% compared with 18.75% in matched RTR-C), giving a relative risk of 13.98. We found statistically significant differences in the frequency of a number of HLA antigens on comparing RTR+C with healthy controls, but none of these differences were found when we compared RTR+C against matched RTR-C.

1. Nicotine (1-100 microM) stimulated both basal and electrically evoked release of 3H-norepinephrine and also caused a transient inhibition of contractions in an in vitro preparation of mouse uterine horns. 2. The inhibitory effect of nicotine on electrically evoked contractions was potentiated by aminophylline (89 micrograms/ml), and overcome by both propranolol (1 microM) and by omitting magnesium from the physiological solution. Acetylcholine (10 microM), in the presence of atropine (10 microM) was able to reproduce the inhibitory effect of nicotine. 3. These pharmacological findings suggest that the inhibitory action of nicotine on electrically evoked contractions in mouse uterus could be indirect, i.e. mediated through the action of this compound on presynaptic nicotine receptors located on adrenergic terminals.

Germinal mosaicism is reported in three siblings with Crouzon syndrome born to normal, unrelated parents. Other probable explanations for the features in this family are discussed.

Using the polymerase chain reaction we have isolated and sequenced cDNA clones corresponding to the polymorphic first domain of the DR beta 1 chain from the DR4, "Dw13" cell line, JHa. We have found that the JHa DR beta 1 allele differs from previously reported Dw13 alleles by a single amino acid substitution at position 86. The functional relevance of this polymorphism is supported by the reactivity pattern of a T-cell clone, E38. E38 is an alloreactive T-cell clone which reacts with all Dw14 stimulator cells and all Dw13-positive cells tested except the "Dw13"-positive homozygous typing cell line JHa. Inhibition studies with monoclonal antibodies revealed the stimulating determinant to be on DR and not on DQ or DP molecules. These data indicate that position 86 of the DR beta 1 chain can play an important role in the formation of determinants recognized by T cells.

The severe Duchenne type of muscular dystrophy is inherited as an X-linked recessive trait. Approximately two thirds of healthy female heterozygous carriers have a high serum creatine kinase (SCK). A suspected carrier with a normal SCK level therefore, presents an important problem in genetic counselling. Based on Bayesian methods, Emery and Hollyway derived a formula which is applicable when the sporadic case is either the son or brother of a consultant and which also includes information on SCK levels in the consultant and in normal daughters and sisters. The present paper describes the results obtained with use of this formula in 27 families with at least a propositus with Duchenne muscular dystrophy.

We investigated the effect of gamma interferon and phorbol ester (TPA), on the expression of HLA class II molecules of myeloid leukaemic cell lines K562, U937, KG-1, HL-60 and ML-2. Gamma interferon induced the expression of HLA-DR but not HLA-DQ on HL-60 and ML-2, increased the expression of HLA-DR and DQ on U937 and induced the expression of HLA-DQ on KG-1. TPA treatment did not affect the expression of HLA class II antigens on U937 and KG-1 and induced the expression of HLA-DR and HLA-DQ on HL-60 and ML-2. TPA treatment did not affect the HLA phenotype of K562 but gamma interferon did induce HLA class I molecules. Thus, gamma interferon cannot only increase the expression of HLA products already expressed on the cells but can also induce the de novo synthesis of these molecules on myeloid leukaemic cell lines.

The HLA class I and class II phenotypes of a panel of 114 unrelated Nigerians have been determined. The panel was tested for all the known class I antigens and comparisons of the HLA-A and -B frequencies with those of other African Negroid populations revealed some differences. Only limited comparisons could be made for the HLA-DR and -D frequencies as these are not available for any well-defined African Negroid population. The data concerning the class II antigens of this panel are the most interesting. Half of the DRw11-positive panel members are DQw3 negative and DQw1 positive. In addition, there is dissociation of some HLA-D and -DR specificities, a number of panel members are positive for an HLA-D specificity and are negative for the corresponding HLA-DR specificity. Our results show the value of population studies in the investigation of the relationship between the different HLA class II antigens.

Several lines of evidence indicate that HLA-Dw, as defined by HTC typing, is not the result of the combined stimulatory effect of HLA-DR and DQ. Therefore, responder cells do not have to share HLA-DQ antigens with the stimulator HTCs to give a typing response. The common HLA-DR-DQ associations observed in HTCs correspond to different patterns of linkage disequilibrium in different populations. HLA-DQ and HLA-Dw are functionally heterogeneous. Although HLA-DQ molecules may play a role in primary stimulation, this role is distinct from that of Dw determinants which have strong lymphocyte activating properties. The role of the HLA-DQ determinants on the other hand, is one of modulating the total T cell response by controlling the proliferation of suppressor and cytotoxic cells. The primary MLC response is the result of the proliferative effect of HLA-Dw, DR, DP, and other associated determinants, in conjunction with a modulatory effect of DQ molecules. However, HLA-Dw (as detected by HTC typing) are DR associated determinants which are immunodominant in primary MLR. The genes of the HLA-DR subregion have been named DR by the WHO nomenclature committee. This subregion encodes the HLA-DR specificities and the DRw52 and DRw53 determinants. Unfortunately this nomenclature does not take into account the need to define the genetic basis of the HLA-Dw determinants--whether they are encoded by separate genes within the HLA-DR subregion or whether they are encoded by as yet unspecified genes in the HLA class II region in linkage disequilibrium with HLA-DR DRw52/53. There are at least three and possibly four beta chain genes in the HLA-DR subregion, all in strong linkage disequilibrium with each other. Some of these are expressed in most haplotypes while others are not; some behave as pseudogenes in some haplotypes and in others, all the genes are expressed. All the genes of the class II region have not been fully characterized. HLA-Dw determinants may be specified by one or more of these genes. When more information becomes available, the genetic and molecular basis of the HLA-Dw series as well as the functional heterogeneity and antigenic strength of the various class II determinants will be better understood.

Deliberate inhalation of volatile substances is a common and harmful practice among young persons worldwide. Recently, we described chromosome damage in children who chronically inhale volatile agents. Clinical and cytogenetic studies were performed for 15 "sniffing" children (13 boys and 2 girls), the purpose of which was to define the chromosomal effect of the acute intoxication. A significant increase in the rate of chromosome abnormalities and in the frequency of sister chromatid exchanges (SCEs) was found in sniffers vs. controls. The values were also higher in children who were acutely intoxicated than in those who chronically inhaled volatile agents. Clinical, socioeconomic, and cytogenetic findings are also discussed.

Forty two white patients of British or Irish descent with coeliac disease and 28 with dermatitis herpetiformis were typed for class I HLA-A, B, and C, and class II DR and DQ antigens. In coeliac disease there was a significant increase in the frequencies of A1, B8, DR3, DR7, and DQw2 compared with controls but no increase of DR2. In dermatitis herpetiformis there were similarly increased frequencies of A1, B8, DR3, and DQw2. In contrast with coeliac disease, however, the frequency of DR7 (18%) was no different from the control group but there was an increased frequency of DR2.

The present paper describes two patients with Sturge-Weber and Klippel Trénaunay-Weber syndromes. Some etiopathogenic factors are analyzed. We suggest that the association of both diseases in the same patient may be due to a single autosomal dominant gene.

We have examined the HLA class II antigenic profiles on different types of leukemic cells and have attempted to relate these findings to the normal differentiation pathways of the cells from which they have arisen. Monoclonal antibodies reacting with the different HLA class II determinants, HLA-DR, DRw52(MT), and DQ, were used to study the expression of these antigens on Epstein-Barr virus transformed cell lines, chronic lymphocytic leukemic cells, acute lymphoblastic leukemic blasts, acute myeloblastic leukemic blasts, and established leukemic cell lines by indirect immunofluorescence binding and immunoprecipitations. The results showed that whereas the HLA-DR and HLA-DRw52(MT2) antigens are normally expressed on the majority of the cells tested, there is a different expression of the HLA-DQ antigens on acute leukemic blasts, chronic lymphocytic leukemic cells, and leukemic cell lines indicating that the DQ molecules may be differentiation antigens preferentially expressed on mature cells. Furthermore, when the pre-B cell leukemic line NALM 6 was induced to differentiate with phorbol ester (TPA), normal expression of the HLA-DQ antigen was obtained after 5 days of culture. The absence of HLA-DQ antigens from the acute leukemic blasts suggests that these immature cells "froze" in the early stages of cell differentiation. We discuss these findings in relation to the role of these HLA class II antigens in cell differentiation and the immune response.

In mice, thymus-derived lymphocytes are differentiated into functional subclasses by their cell surface antigens. The Ly 1 determinants are present on T cells with a helper function, whereas Ly 2 and Ly 3 antigens are expressed on the surface of lymphocytes with suppressor or cytotoxic functions. In man also, T-cell subsets have been identified using allo- and heteroimmune sera and, more recently, using monoclonal antibodies, which seem to identify helper and suppressor or cytotoxic subpopulations. The major histocompatibility system (MHS)-encoded Ia antigens belong to several polymorphic families of membrane associated glycoproteins originally found on B lymphocytes; however, they have also been shown to be markers for suppressor T cells in mice. Recent studies have shown that in both mouse and man, T cells activated by a mixed lymphocyte reaction or by mitogens become Ia+. Furthermore, some human T lymphoid cells, either freshly isolated from peripheral blood or after in vitro activation by lectins or alloantigens, possess suppressor properties. We report here the phenotype of a T suppressor-cell subpopulation which was induced in long-term culture of lymphoid cells after activation with phytohaemagglutinin (PHA). Our results suggest that a subset of T cells was progressively expanded over a period of 8 days in culture and that, with the expression on the surface of these cells of 'Ia-like' antigens, they acquired the capacity to suppress the proliferative response of syngeneic or allogeneic lymphocytes to alloantigens or mitogens.

The present paper describes a homozygote patient for the gene of hemoglobin S. The patient is the first child of non-consanguineous parents. The family study revealed 11 heterozygote subjects for the sickle cell disease.

HLA class II antigens were studied in a panel of 130 Nigerians. Complex patterns of associations were seen between HLA-Dw, -DR and -DQ specificities, differing widely from those reported for other populations. A number of Dw types were associated with the same DR antigen: Dw'1N' and Dw'BERN' with DR1, Dw2 and Dw'2N' with DR2, Dw5 and Dw'5N' (Dw5 + Dw'F5') with DRw11. It was also observed that a Dw type associated with more than one DR antigen: HLA-Dw3 was assigned to individuals who were DR3 negative and similarly Dw10, Dw13 and Dw14 to individuals negative for DR4. HLA-DRw8 and Dw8 were completely dissociated in Nigerians, and Dw8 did not show a preferential DR association. These results demonstrate that DR and DQ identity between HTC stimulator and responder cell is not necessarily a prerequisite for Dw to be assigned. Preliminary studies show that subtypes of HLA-Dw1 and Dw8 detected by HTC typing correlate with restriction fragment length polymorphisms (RFLPs) detected with a combination of Bgl II enzyme and DRA/DRB cDNA probes. HLA-DP antigen frequencies differed between Nigerians and British Caucasoids. The most common DP antigen in Nigerians was DPw1, compared with DPw4 in Caucasoids. HLA-DPw6 appeared to be absent or rare in both Nigerians and British Caucasoids. Only five out of 68 Nigerians tested were assigned two DP specificities. The association between HLA-DR3 and DPw1 reported in Caucasoid panels was absent in Nigerians.