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1.  Precautions surrounding blood transfusion in autoimmune haemolytic anaemias are overestimated 
Blood Transfusion  2015;13(4):616-621.
It is very evident that many precautions are taken regarding transfusion of red blood cells in patients with autoimmune haemolytic anaemia. Frequently, considerable efforts are made to examine the indication and serological compatibility prior to transfusion in such patients. However, at times, this may unnecessarily jeopardize patients who urgently require a red blood cell transfusion.
Materials and methods
Thirty-six patients with warm-type autoimmune haemolytic anaemia were included in this study. All patients had reactive serum autoantibodies and required blood transfusion. Standard serological assays were employed for the detection and characterization of antibodies to red blood cells.
A positive direct antiglobulin test was observed in all 36 patients, in addition to detectable antibodies in both the eluate and serum. Significant alloantibodies were detected in the serum samples of three patients (anti-c, anti-JKa, and anti-E). In 32 patients, red blood cell transfusion was administered with no significant haemolytic transfusion reactions due to auto- and/or allo-antibodies. Due to overestimation of positive cross-matches three patients received no transfusion or delayed transfusion and died, and one patient died due to unrecognised blood loss and anaemia which was attributed to an ineffective red blood cell transfusion.
Many of the reported recommendations regarding transfusion of red blood cells in autoimmune haemolytic anaemia are highly questionable, and positive serological cross-matches should not result in a delay or refusal of necessary blood transfusions.
PMCID: PMC4624538  PMID: 26192772
positive cross-match; serological incompatibility; incompatible RBC transfusion; autoimmune haemolytic anaemia; alloantibodies
2.  Comparative Biomechanical and Microstructural Analysis of Native versus Peracetic Acid-Ethanol Treated Cancellous Bone Graft 
BioMed Research International  2014;2014:784702.
Bone transplantation is frequently used for the treatment of large osseous defects. The availability of autologous bone grafts as the current biological gold standard is limited and there is a risk of donor site morbidity. Allogenic bone grafts are an appealing alternative, but disinfection should be considered to reduce transmission of infection disorders. Peracetic acid-ethanol (PE) treatment has been proven reliable and effective for disinfection of human bone allografts. The purpose of this study was to evaluate the effects of PE treatment on the biomechanical properties and microstructure of cancellous bone grafts (CBG). Forty-eight human CBG cylinders were either treated by PE or frozen at −20°C and subjected to compression testing and histological and scanning electron microscopy (SEM) analysis. The levels of compressive strength, stiffness (Young's modulus), and fracture energy were significantly decreased upon PE treatment by 54%, 59%, and 36%, respectively. Furthermore, PE-treated CBG demonstrated a 42% increase in ultimate strain. SEM revealed a modified microstructure of CBG with an exposed collagen fiber network after PE treatment. We conclude that the observed reduced compressive strength and reduced stiffness may be beneficial during tissue remodeling thereby explaining the excellent clinical performance of PE-treated CBG.
PMCID: PMC3942278  PMID: 24678514
3.  Advanced Therapy Medicinal Products – a Multiple Challenge 
PMCID: PMC3901633  PMID: 24474887
4.  Whither Advanced Therapy Medicinal Products? 
PMCID: PMC3901618  PMID: 24474896
5.  Validation of Serological Testing for Anti-Treponema pallidum from Postmortem Blood on the Siemens-BEP®-III Automatic System 
Infectious disease marker testing is obligatory for the release of human tissue for transplantation. Most CE-marked tests are not validated for postmortem blood. In a previous study we have validated the testing for anti-HIV-1/2, anti-HCV, HBsAg, and anti-HBc. Here, we present the validation of testing for antibodies against T. pallidum, which is the last marker obligatory for tissue release for transplantation.
17 samples of postmortem sera and 10 samples of both pre-und postmortem sera were obtained from cornea donors and tested for anti-T. pallidum on the Siemens-BEP-III-System. These sera were spiked with anti-T. pallidum-positive standard sera in concentrations which give low- and high-positive results at the respective dilution.
Two of the unspiked postmortem sera were false-positive most likely due to intense hemolysis (free hemoglobin > 50 mg/dl). Of the 25 negative postmortem sera, none of the spiked samples was false-negative after 0, 24 and 60 h.
There is no indication that postmortem samples give false-negative or false-positive results with the test system and test kits used in cases of low hemolysis. The procedure described might serve as a model for validating other test kits on postmortem samples.
PMCID: PMC3901593  PMID: 24474889
Infectious disease serology; Postmortem blood; Blood testing; Tissue donation; Treponema pallidum
6.  Six years of clinical follow-up with endothelial cell–seeded small-diameter vascular grafts during coronary bypass surgery 
Journal of Tissue Engineering  2013;4:2041731413504777.
This clinical study was performed to investigate the patency rate of endothelial cell–seeded small-diameter expanded polytetrafluoroethylene grafts during coronary artery bypass surgery. Between September 1995 and December 1998, 14 patients (median age: 71 years, range: 61–79 years) received 21 endothelial cell–seeded small-diameter grafts. In all, 43% of the performed implantations were reoperations. Endothelial cells were harvested from a forearm vein, cultured and characterized in the laboratory until a sufficient number was available. After in vitro seeding, the grafts were allowed to mature for another 10 days, prior to implantation. Graft patency was investigated with angiography, angioscopy, and intravascular ultrasonography during follow-up. Cumulative data represented 58 patients’ years and was 100% complete. The seeded autologous vascular endothelial cell density was 1.05 × 105 ± 0.12 × 105 cells/cm2 with a cell viability of 95.5 ± 1.5%. Operative mortality was 7.1% (one patient). Patency rate at discharge was 95.2%, and at a mean follow-up of 27 months was 90.5%. The proven patency rate at up to 72 months was at least 50.0%, as five patients refused angiographic evaluation. None of these five patients suffered from angina pectoris and so the best scenario would have shown a patency rate of 85.7%. Angioscopy and intravascular ultrasonography showed absence of atheroma or stenosis in the investigated patent grafts. Autologous vascular endothelial cell seeding improves patency rate of small-caliber expanded polytetrafluoroethylene grafts in patients without suitable autologous graft material.
PMCID: PMC3764981  PMID: 24020013
Polytetrafluoroethylene (PTFE) grafts; endothelial cells; in vitro seeding; alternative bypass material
7.  Current Trends in Tissue Banking 
PMCID: PMC3678254  PMID: 23801929
8.  Comparison of in situ Corneoscleral Disc Excision versus Whole Globe Enucleation in Cornea Donors Regarding Microbial Contamination in Organ Culture Medium – a Prospective Monocentric Study over 9 Years 
Corneas needed for keratoplasty can be harvested using two techniques: whole globe enucleation and in situ excision of the corneoscleral disc. This study evaluates the rate of microbial contamination of the donor cornea organ culture medium according to the method of retrieval.
All donor corneas of our cornea bank received between January 1, 2001 and December 31, 2009 put into organ culture and microbio-logically tested were prospectively analyzed for microbial contamination of the organ culture medium.
2,805 donor corneas could be included in this study in total. 975 of them were retrieved by whole globe enucleation (group 1) and 1,830 by in situ corneoscleral disc excision (group 2). 15 corneas of group 1 (1.5%) and 46 corneas of group 2 (2.5%) showed a contamination of the organ culture medium. The difference was shown not to be statistically significant (p = 0.082).
The rate of microbial contamination in organ-cultured donor corneas does not seem to be dependent on the method of their retrieval.
PMCID: PMC3678256  PMID: 23801381
Cornea donation; Whole globe enucleation; In situ corneoscleral disc excision; Organ culture; Microbiology
9.  Validation of the Microbiological Testing of Tissue Preparations Using the BACTEC™ Blood Culture System 
Since blood culture bottles are validated by the manufacturer for blood only, an additional validation for the use with fluids of tissue preparations is necessary.
Two 10-ml samples of cornea culture medium, histidine-tryptophan-ketoglutarate (HTK) solution, or Ringer solution at the end of femur head thermo-disinfection were given into blood culture bottles (BD BACTEC™ Plus Aerobic/F, Anaerobic/F for cornea culture medium and BD BACTEC™ Standard Aerobic/Anaerobic for HTK and Ringer solution) and subsequently spiked with 10–100 colony forming units (CFU) of bacteria or fungi (aerobic bacteria: Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa; anaerobic bacteria: Clostridium sporogenes; fungi: Candida albicans, Aspergillus brasiliensis) according to the European Pharmacopoeia Chapter 2.6.1. Results: All tested bacteria and fungi could be detected in all solutions. All positive and negative controls were tested correctly. Compared to the positive controls, the microbial growth was delayed in the antibiotic-containing cornea culture medium, and negative in two cases of B. subtilis spiking. Conclusion: The use of BACTEC™ blood culture bottles seems to be a suitable method for microbiological testing of HTK solution, Ringer solution, and, with limitations, also for testing of the antibiotic-containing cornea culture medium.
PMCID: PMC3678268  PMID: 23801426
Validation; Microbiology; Tissue
10.  Virus NAT for HIV, HBV, and HCV in Post-Mortal Blood Specimens over 48 h after Death of Infected Patients – First Results 
According to EU regulations (EU directive 2006/17/EC), blood specimens for virologic testing in the context of post-mortal tissue donation must be taken not later than 24 h post mortem.
To verify validity of NAT in blood specimens collected later, viral nucleic acid concentrations were monitored in blood samples of deceased persons infected with HIV (n = 7), HBV (n = 5), and HCV (n = 17) taken upon admission and at 12 h, 24 h, 36 h and 48 h post mortem. HIV and HCV RNA were quantified using Cobas TaqMan (Roche), HBV DNA was measured by in-house PCR.
A more than 10-fold decrease of viral load in samples taken 36 h or 48 h post mortem was seen in one HIV-infected patient only. For all other patients tested the decrease of viral load in 36-hour or 48-hour post-mortal samples was less pronounced. Specimens of 3 HIV- and 2 HBV-infected patients taken 24 h post mortem or later were even found to have increased concentrations (>10-fold), possibly due to post-mortem liberation of virus from particular cells or tissues.
Our preliminary data indicate that the time point of blood collection for HIV, HBV and HCV testing by PCR may be extended to 36 h or even 48 h post mortem and thus improve availability of tissue donations.
PMCID: PMC3678255  PMID: 23801336
Post-mortal tissue donation; Virus safety NAT screening
11.  Coding of Tissue Preparations with Eurocode in Germany 
A safe look back of products requires their unique identification. Blood products are encoded in Germany with Eurocode since 1987. EU Directives 2004/23/EC und 2006/86/EC demanded unique identification and safe look back procedure also for tissues and cells. Eurocode IBLS e.V. and the DGTI working parties ‘Tissue Preparations’ and ‘Automation and Data Processing’ supplemented the already available Eurocode nomenclature for blood products with further data structures for tissue preparations and deliberated the federal authorities during the EU hearings. In result all EU member states can administer the coding system oneself, but have to take care about the ‘key code’ structure as defined and the common part at the begin of the ID number of the preparations. Eurocode today offers an EU-conform coding system considering various aspects of blood, tissue and cell preparations in an ISO-standardized form.
PMCID: PMC3678294  PMID: 23801930
Eurocode; Tissue; Coding systems; Look back; European Directive 2006/86/EC
12.  Thawed solvent/detergent-treated plasma: too precious to be wasted after 6 hours? 
Blood Transfusion  2012;10(3):360-367.
Coagulopathy associated with trauma and bleeding requires early administration of haemostatic agents. Solvent/detergent-treated plasma (S/D-plasma) requires thawing and its availability for clinical use is, therefore, delayed. The long-term stability of clotting factors in thawed S/D-plasma has not been thoroughly investigated. The purpose of this study was to evaluate stability of clotting factors and inhibitors in thawed S/D-plasma stored at 4 °C for 6 days.
Materials and methods.
Clotting factor levels and bacterial contamination were investigated using 20 units of S/D-plasma. Fibrinogen, factor (F) II, FV, FVII, FVIII, FIX, FX, FXI, FXII, FXIII, antithrombin, von Willebrand antigen (VWF-Ag), plasmin inhibitor, protein C and free protein S were analysed over time.
After 6 days of storage the results were as follows: fibrinogen 270 mg/dL (−10 mg/dL, p=0.0204), FII 75% (−5%, p<0.0001), FV 88% (−14%, p<0.0001), FVII 81% (−24%, p<0.0001), FVIII 70% (−16%, p<0.0001), FIX 96% (−8, p<0.0001), FX 92% (−1%, p<0.0001), FXI 119% (−4%, p=0.3666), FXII 94% (−2%, p=0.3602), FXIII 89% (−1%, p 0.0019), free protein S 76% (−4%, p<0.0001), protein C 96% (+1%, p=0.0371), antithrombin 92% (−3%, p<0.0001), plasmin inhibitor 29% (−4%, p<0.0299), VWF-Ag 137% (+2%, p=0.2205). FVII and FVIII showed a critical drop of more than 20% or approached the lower quality assurance threshold after storage for more than 24 hours. No S/D-plasma showed bacterial contamination.
All clotting factors in thawed S/D plasma remained stable for up to 24 hours when stored at 4 °C. Storage of thawed S/D plasma may improve the availability of this product in emergency situations.
PMCID: PMC3417736  PMID: 22507858
S/D-plasma; massive transfusion; plasma storage; clotting factors
13.  Inactivation Effect of Standard and Fractionated Electron Beam Irradiation on Enveloped and Non-Enveloped Viruses in a Tendon Transplant Model 
For increasing allograft tendon safety in reconstructive surgery, an effective sterilization method achieving sterility assurance including viruses without impairing the grafts properties is needed. Fractionated Electron Beam (Ebeam) has shown promising in vitro results. The proof of sufficient virus inactivation is a central part of the process validation.
The Ebeam irradiation of the investigated viruses was performed in an optimized manner (oxygen content < 0.1%, −78 °C). Using principles of a tendon model the virus inactivation kinetics for HIV-2, HAV, pseudorabies virus (PRV) and porcine parvovirus (PPV) were calculated as TCID50/ml and D10 value (kGy) for the fractionated (10 × 3.4 kGy) and the standard (1 × 34 kGy) Ebeam irradiation.
All viruses showed comparable D10 values for both Ebeam treatments. For sufficient virus titer reduction of 4 log10 TCID50/ml, a dose of 34 kGy of the fractionated Ebeam irradiation was necessary in case of HIV-2, which was the most resistant virus investigated in this study.
The fractionated and the standard Ebeam irradiation procedure revealed comparable and sufficient virus inactivation capacities. In combination with the known good biomechanical properties of fractionated Ebeam irradiated tendons, this method could be a safe and effective option for the terminal sterilization of soft tissue allografts.
PMCID: PMC3388620  PMID: 22896764
Allograft; Irradiation; Ebeam; Tissue sterilization; Tissue bank
14.  Validation of the Serological Testing for Anti-HIV-1/2, Anti-HCV, HBsAg, and Anti-HBc from Post-mortem Blood on the Siemens-BEP-III Automatic System 
Some properties of blood are modified post mortem. These modifications might give false-negative or false-positive results in infectious disease testing. Most CE-marked test equipment for infectious serology testing is not validated for testing post-mortal blood. Validation, however, is obligatory, if the results are used for the release of tissues for transplantation.
Samples of pre- and post-mortem sera were obtained from 20 cornea donors, and the results were compared for anti-HIV-1/2, anti-HCV, HBsAg, and anti-HBc on the Siemens-BEP-III Automatic System. Negative post-mortem sera were spiked with standard sera (PEI anti-HCV IgG, PEI HBsAg ad 1000 standard, anti-HBc IgG (WHO) NIBSC 95/522, PEI anti-HIV-IV) in concentrations which give low- and high-positive results for the respective marker.
All pre-mortem sera were negative for all markers. None of the post-mortem samples was false-positive. None of the spiked postmortem samples was false-negative. Technical errors occurred during the validation process but could be detected and eliminated. Serum samples should be centrifuged immediately after collection, and it must be taken into account that post-mortem serum could rarely lead to blockage of pipetting systems due to clotting phenomena.
There is no indication that post-mortem samples give false-negative or false-positve results with the test system and test kits used. The procedure described might serve as a model for validating other test kits on post-mortem samples.
PMCID: PMC3267999  PMID: 22403520
Infectious disease serology·; Post-mortem blood·; Tissue donation·; BEP-III·Validation
15.  Allogenic versus autologous cancellous bone in lumbar segmental spondylodesis: a randomized prospective study 
European Spine Journal  2009;18(5):687-695.
The current gold standard in lumbar fusion consists of transpedicular fixation in combination with an interbody interponate of autologous bone from iliac crest. Because of the limited availability of autologous bone as well as the still relevant donor site morbidity after iliac crest grafting the need exists for alternative grafts with a comparable outcome. Forty patients with degenerative spinal disease were treated with a monosegmental spondylodesis (ventrally, 1 PEEK-cage; dorsally, a screw and rod system), and randomly placed in two groups. In group 1, autogenous iliac crest cancellous bone was used as a cage filling. In group 2 the cages were filled with an allogenic cancellous bone graft. Following 3, 6, 9 and 12 months, the clinical outcome was determined on the basis of: the Oswestry Low Back Pain Disability Questionnaire; patient satisfaction; patient willingness to undergo the operation again; and a visual analog scale for pain. The radiological outcome was based on both fusion rate (radiographs, computed tomography), and on the bone mineral density of the grafts. After 6 months, the X-rays of the patients in group 2 had a significantly lower rate of fusion. Aside from this, there were no further significant differences. After 12 months, radiological results showed a similar fusion rate in both groups. Donor site complications consisted of five patients with hematoma, and three patients with persistent pain in group 1. No implant complications were observed. If a bone bank is available for support and accepting the low risk of possible transmission of infectious diseases, freeze–dried allogenic cancellous bone can be used for monosegmental spondylodeses. The results demonstrated an equivalent clinical outcome, as well as similar fusion rates following a 12-month period. This is in despite of a delayed consolidation process.
PMCID: PMC3234005  PMID: 19148687
Circumferential lumbar fusion; Allogenic cancellous bone; Autologous bone; PEEK-cage; Bone grafts
16.  Remodeling of ACL Allografts is Inhibited by Peracetic Acid Sterilization 
Sterilization of allografts for anterior cruciate ligament (ACL) reconstruction has become an important prerequisite to prevent disease transmission. However, current sterilization techniques impair the biological or mechanical properties of such treated grafts. Peracetic acid (PAA) has been successfully used to sterilize bone allografts without these disadvantages and does not impair the mechanical properties of soft tissue grafts in vitro. We asked whether PAA sterilization would influence recellularization, restoration of crimp length and pattern, and revascularization of ACL grafts during early healing. We used an in vivo sheep model for open ACL reconstruction. We also correlated the histologic findings with the restoration of anteroposterior stability and structural properties during load-to-failure testing. PAA slowed remodeling activity at 6 and 12 weeks compared to nonsterilized allografts and autografts. The mechanical properties of PAA grafts were also reduced compared to these control groups at both time points. We conclude PAA sterilization currently should not be used to sterilize soft tissue grafts typically used in ACL reconstruction.
PMCID: PMC2584264  PMID: 18491201
17.  CCR5Δ32 Genotypes in a German HIV-1 Seroconverter Cohort and Report of HIV-1 Infection in a CCR5Δ32 Homozygous Individual 
PLoS ONE  2008;3(7):e2747.
Homozygosity (Δ32/Δ32) for the 32 bp deletion in the chemokine receptor 5 (CCR5) gene is associated with strong resistance against HIV infection. Heterozygosity is associated with protection of HIV-1 disease progression.
Methodology/Principal Findings
We genotyped a population of 737 HIV-positive adults and 463 healthy controls for the CCR5Δ32 deletion and found heterozygous frequencies of 16.2% (HIV-negative) and 17.5% (HIV-positive) among Caucasian individuals. Analysis of CCR5Δ32 influence on disease progression showed notably lower viral setpoints and a longer time to a CD4 count of <200 µl−1 in seroconverters heterozygous for the deletion. Furthermore, we identified one HIV-positive man homozygous for the Δ32 deletion.
The protective effect of CCR5 Δ32 heterozygosity is confimed in a large cohort of German seroconverters. The HIV-infected CCR5 Δ32 homozygous individual, however, displays extremely rapid disease progression. This is the 12th case of HIV-infection in this genotype described worldwide.
PMCID: PMC2453227  PMID: 18648518
18.  Decrease in expression of bone morphogenetic proteins 4 and 5 in synovial tissue of patients with osteoarthritis and rheumatoid arthritis 
Bone morphogenetic proteins (BMPs) have been identified as important morphogens with pleiotropic functions in regulating the development, homeostasis and repair of various tissues. The aim of this study was to characterize the expression of BMPs in synovial tissues under normal and arthritic conditions. Synovial tissue from normal donors (ND) and from patients with osteoarthritis (OA) and rheumatoid arthritis (RA) were analyzed for BMP expression by using microarray hybridization. Differential expression of BMP-4 and BMP-5 was validated by semiquantitative RT-PCR, in situ hybridization and immunohistochemistry. Activity of arthritis was determined by routine parameters for systemic inflammation, by histological scoring of synovitis and by semiquantitative RT-PCR of IL-1β, TNF-α, stromelysin and collagenase I in synovial tissue. Expression of BMP-4 and BMP-5 mRNA was found to be significantly decreased in synovial tissue of patients with RA in comparison with ND by microarray analysis (p < 0.0083 and p < 0.0091). Validation by PCR confirmed these data in RA (p < 0.002) and also revealed a significant decrease in BMP-4 and BMP-5 expression in OA compared with ND (p < 0.015). Furthermore, histomorphological distribution of both morphogens as determined by in situ hybridization and immunohistochemistry showed a dominance in the lining layer of normal tissues, whereas chronically inflamed tissue from patients with RA revealed BMP expression mainly scattered across deeper layers. In OA, these changes were less pronounced with variable distribution of BMPs in the lining and sublining layer. BMP-4 and BMP-5 are expressed in normal synovial tissue and were found decreased in OA and RA. This may suggest a role of distinct BMPs in joint homeostasis that is disturbed in inflammatory and degenerative joint diseases. In comparison with previous reports, these data underline the complex impact of these factors on homeostasis and remodeling in joint physiology and pathology.
PMCID: PMC1526630  PMID: 16542506
19.  Analysis of immunoglobulin light chain rearrangements in the salivary gland and blood of a patient with Sjögren's syndrome 
Arthritis Research  2002;4(4):R4.
Patients with Sjögren's syndrome (SS) have characteristic lymphocytic infiltrates of the salivary glands. To determine whether the B cells accumulating in the salivary glands of SS patients represent a distinct population and to delineate their potential immunopathologic impact, individual B cells obtained from the parotid gland and from the peripheral blood were analyzed for immunglobulin light chain gene rearrangements by PCR amplification of genomic DNA. The productive immunglobulin light chain repertoire in the parotid gland of the SS patient was found to be restricted, showing a preferential usage of particular variable lambda chain genes (Vλ2E) and variable kappa chain genes (VκA27). Moreover, clonally related VL chain rearrangements were identified; namely, VκA27–Jκ5 and VκA19–Jκ2 in the parotid gland, and Vλ1C–Jλ3 in the parotid gland and the peripheral blood. Vκ and Vλ rearrangements from the parotid gland exhibited a significantly elevated mutational frequency compared with those from the peripheral blood (P < 0.001). Mutational analysis revealed a pattern of somatic hypermutation similar to that found in normal donors, and a comparable impact of selection of mutated rearrangements in both the peripheral blood and the parotid gland. These data indicate that there is biased usage of VL chain genes caused by selection and clonal expansion of B cells expressing particular VL genes. In addition, the data document an accumulation of B cells bearing mutated VL gene rearrangements within the parotid gland of the SS patient. These results suggest a role of antigen-activated and selected B cells in the local autoimmune process in SS.
PMCID: PMC125296  PMID: 12106503
B cells; parotid gland; Sjögren's syndrome; somatic mutation; V light chain genes
20.  Perturbations in the impact of mutational activity on Vλ genes in systemic lupus erythematosus 
Arthritis Research  2001;3(6):368-374.
To assess the impact of somatic hypermutation and selective influences on the Vλ light chain repertoire in systemic lupus erythematosus (SLE), the frequency and pattern of mutations were analyzed in individual CD19+ B cells from a patient with previously undiagnosed SLE. The mutational frequency of nonproductive and productive rearrangements in the SLE patient was greater (3.1 × 10-2 vs 3.4 × 10-2, respectively) than that in normal B cells (1.2 × 10-2 vs 2.0 × 10-2, both P < 0.001). The frequencies of mutated rearrangements in both the nonproductive and productive repertoires were significantly higher in the patient with SLE than in normal subjects. Notably, there were no differences in the ratio of replacement to silent (R/S) mutations in the productive and nonproductive repertoires of the SLE patient, whereas the R/S ratio in the framework regions of productive rearrangements of normal subjects was reduced, consistent with active elimination of replacement mutations in this region. The pattern of mutations was abnormal in the SLE patient, with a significant increase in the frequency of G mutations in both the productive and nonproductive repertoires. As in normal subjects, however, mutations were found frequently in specific nucleotide motifs, the RGYW/WRCY sequences, accounting for 34% (nonproductive) and 46% (productive) of all mutations. These data are most consistent with the conclusion that in this SLE patient, the mutational activity was markedly greater than in normal subjects and exhibited some abnormal features. In addition, there was decreased subsequent positive or negative selection of mutations. The enhanced and abnormal mutational activity along with disturbances in selection may play a role in the emergence of autoreactivity in this patient with SLE.
PMCID: PMC64848  PMID: 11714391
autoimmunity; B cells; SLE; somatic hypermutation; V genes

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