Timing of radiotherapy (RT) after radical prostatectomy (RP): long-term outcomes in the RADICALS-RT trial (NCT00541047).

BACKGROUND: The optimal timing of radiotherapy (RT) after radical prostatectomy for prostate cancer has been uncertain. RADICALS-RT compared efficacy and safety of adjuvant RT versus an observation policy with salvage RT for prostate-specific antigen (PSA) failure. PATIENTS AND METHODS: RADICALS-RT was a randomised controlled trial enrolling patients with ≥1 risk factor (pT3/4, Gleason 7-10, positive margins, preoperative PSA≥10 ng/ml) for recurrence after radical prostatectomy. Patients were randomised 1:1 to adjuvant RT ('Adjuvant-RT') or an observation policy with salvage RT for PSA failure ('Salvage-RT') defined as PSA≥0.1 ng/ml or three consecutive rises. Stratification factors were Gleason score, margin status, planned RT schedule (52.5 Gy/20 fractions or 66 Gy/33 fractions) and treatment centre. The primary outcome measure was freedom-from-distant-metastasis (FFDM), designed with 80% power to detect an improvement from 90% with Salvage-RT (control) to 95% at 10 years with Adjuvant-RT. Secondary outcome measures were biochemical progression-free survival, freedom from non-protocol hormone therapy, safety and patient-reported outcomes. Standard survival analysis methods were used; hazard ratio (HR)<1 favours Adjuvant-RT. RESULTS: Between October 2007 and December 2016, 1396 participants from UK, Denmark, Canada and Ireland were randomised: 699 Salvage-RT, 697 Adjuvant-RT. Allocated groups were balanced with a median age of 65 years. Ninety-three percent (649/697) Adjuvant-RT reported RT within 6 months after randomisation; 39% (270/699) Salvage-RT reported RT during follow-up. Median follow-up was 7.8 years. With 80 distant metastasis events, 10-year FFDM was 93% for Adjuvant-RT and 90% for Salvage-RT: HR=0.68 [95% confidence interval (CI) 0.43-1.07, P=0.095]. Of 109 deaths, 17 were due to prostate cancer. Overall survival was not improved (HR=0.980, 95% CI 0.667-1.440, P=0.917). Adjuvant-RT reported worse urinary and faecal incontinence 1 year after randomisation (P=0.001); faecal incontinence remained significant after 10 years (P=0.017). CONCLUSION: Long-term results from RADICALS-RT confirm adjuvant RT after radical prostatectomy increases the risk of urinary and bowel morbidity, but does not meaningfully improve disease control. An observation policy with salvage RT for PSA failure should be the current standard after radical prostatectomy. TRIAL IDENTIFICATION: RADICALS, RADICALS-RT, ISRCTN40814031, NCT00541047.

Targeting GM-CSF for collagenase-induced osteoarthritis pain and disease in mice.

OBJECTIVES: Pharmacological options for treating osteoarthritis (OA) are limited and alternative treatments are required. Given the clinical data indicating that granulocyte macrophage-colony stimulating factor (GM-CSF) may be a therapeutic target in human OA, we evaluated different treatment regimens with a neutralizing anti-GM-CSF monoclonal antibody (mAb) in an experimental OA model to determine their effectiveness on amelioration of pain and disease. METHODS: The collagenase-induced osteoarthritis (CiOA) model was induced in C57BL/6 mice, followed by different treatment regimens of anti-GM-CSF mAb or isotype control. Anti-CCL17 mAb treatment was also administered continually during the late stage of CiOA. Pain-related behavior (change in weight distribution of hind limbs), and disease (cartilage damage and osteophyte size) were assessed. RESULTS: Blocking GM-CSF only during early synovitis in CiOA prevented pain and disease development. Once OA pain was established, regardless of the treatment regimen, anti-GM-CSF mAb treatment rapidly and efficiently ameliorated it; however, unless the treatment was continued, pain returned and disease progressed. Continual late stage blockade of GM-CSF was able to ameliorate pain (between-group difference: -6.567; 95% confidence interval (CI): -10.12, -3.011) and suppress cartilage damage (P = 0.0317, 95% CI: -1.75, -0.0556). Continual late stage blockade of CCL17 showed similar effects on pain and disease development. CONCLUSIONS: Early and short-term GM-CSF neutralization is effective at preventing CiOA pain and disease development but, once pain is evident, continual GM-CSF blockade is required to prevent pain from returning and to suppress disease progression in mice. These data reinforce the potential benefits of anti-GM-CSF (and anti-CCL17) mAb therapy in OA and should inform further clinical trials.

Pulseless electrical activity following traumatic cardiac arrest: Sign of life or death?

BACKGROUND: Generally considered a sign of life, PEA is the most common arrhythmia encountered following pre-hospital traumatic cardiac arrest. Some recommend cardiac ultrasound (CUS) to determine cardiac wall motion (CWM) prior to terminating resuscitation efforts. This purpose of this study was to evaluate the outcomes of patients with traumatic cardiac arrest presenting with PEA, with and without CWM. METHODS: Trauma patients who underwent pre-hospital CPR were identified from the registries of two level-1 trauma centers. Pre-hospital management by emergency medical transport services was guided by advanced life support protocols. The on-duty trauma surgeon directed the resuscitations and performed or supervised CUS and determined CWM. RESULTS: Among 277 patients who underwent pre-hospital CPR, 110 patients had PEA on arrival to ED. 69 (62.7%) were injured by blunt mechanisms. Median CPR duration was 20.0 and 8.0 min for pre-hospital and ED, respectively. Sixty-three patients (22.7%) underwent resuscitative thoracotomy. One hundred seventy-two patients (62.1%) received CUS and of these 32 (18.6%) had CWM. CWM was significantly associated with survival to hospital admission (21.9% vs. 1.4%; P < 0.001); however, no patient with CUS survived to hospital discharge. Overall, only one patient with PEA on arrival survived to discharge. CONCLUSION: Following pre-hospital traumatic cardiac arrest, PEA on arrival portends death. Although CWM is associated with survival to admission, it is not associated with meaningful survival. Heroic resuscitative measures may be unwarranted for PEA following pre-hospital traumatic arrest, regardless of CWM.

Identification of epistasis through a partial advanced intercross reveals three arthritis loci within the Cia5 QTL in mice.

Identification of genes controlling complex diseases has proven to be difficult; however, animal models may pave the way to determine how low penetrant genes interact to promote disease development. We have dissected the Cia5/Eae3 susceptibility locus on mouse chromosome 3 previously identified to control disease in experimental models of multiple sclerosis and rheumatoid arthritis. Congenic strains showed significant but small effects on severity of both diseases. To improve the penetrance, we have now used a new strategy that defines the genetic interactions. The QTL interacted with another locus on chromosome 15 and a partial advanced intercross breeding of the two congenic strains for eight generations accumulated enough statistical power to identify interactions with several loci on chromosome 15. Thereby, three separate loci within the original QTL could be identified; Cia5 affected the onset of arthritis by an additive interaction with Cia31 on chromosome 15, whereas the Cia21 and Cia22 affected severity during the chronic phase of the disease through an epistatic interaction with Cia32 on chromosome 15. The definition of genetic interactions was a prerequisite to dissect the Cia5 QTL and we suggest the partial advanced intercross strategy to be helpful also for dissecting other QTL controlling complex phenotypes.

Blockade of collagen-induced arthritis post-onset by antibody to granulocyte-macrophage colony-stimulating factor (GM-CSF): requirement for GM-CSF in the effector phase of disease.

There is mounting evidence for a role of the growth factor granulocyte-macrophage colony-stimulating factor (GM-CSF) in inflammatory disease, including arthritis. In the present study, we examined the effectiveness of treatment of collagen-induced arthritis (CIA) with a neutralizing mAb to GM-CSF. DBA/1 mice were immunized for the development of CIA and treated at different times, and with different doses, with neutralizing mAb to GM-CSF or isotype control mAb. Anti-GM-CSF mAb treatment prior to the onset of arthritis, at the time of antigen challenge, was effective at ameliorating the ensuing disease. Modulation of arthritis was seen predominantly as a reduction in overall disease severity, both in terms of the number of limbs affected per mouse and the clinical score of affected limbs. Importantly, anti-GM-CSF mAb treatment ameliorated existing disease, seen both as a reduction in the number of initially affected limbs progressing and lower numbers of additional limbs becoming affected. By histology, both inflammation and cartilage destruction were reduced in anti-GM-CSF-treated mice, and the levels of tumor necrosis factor-a and IL-1beta were also reduced in joint tissue washouts of these mice. Neither humoral nor cellular immunity to type II collagen, however, was affected by anti-GM-CSF mAb treatment. These results suggest that the major effect of GM-CSF in CIA is on mediating the effector phase of the inflammatory reaction to type II collagen. The results also highlight the essential role of GM-CSF in the ongoing development of inflammation and arthritis in CIA, with possible therapeutic implications for rheumatoid arthritis.

Chest radiographs of limited utility in the diagnosis of blunt traumatic aortic laceration.

BACKGROUND: The radiographic diagnosis of blunt traumatic aortic laceration (BTAL) remains problematic. We reviewed our experience with chest radiographic signs of BTAL at a single trauma center. METHODS: The chest radiographs of 188 consecutive blunt trauma patients with suspected BTAL who underwent portable chest radiography and aortography were retrospectively reviewed by a thoracic radiologist. The presence or absence of 15 radiographic findings were recorded, and the sensitivity and specificity of individual radiographic signs and combinations of signs were determined. RESULTS: There were 10 patients with BTAL. Although three signs showed greater than 90% sensitivity for BTAL, these signs showed low specificity, and no significant improvement in overall accuracy was achieved by combining radiographic findings. CONCLUSION: The experience at our institution suggests that chest radiographs have limited utility in the accurate diagnosis of blunt traumatic aortic laceration. Cross-sectional imaging techniques will likely become the preferred imaging procedures for evaluating patients with suspected BTAL.

Antibodies to type II collagen and HLA disease susceptibility markers in rheumatoid arthritis.

OBJECTIVE: To seek associations between antibodies to native and denatured type II collagen (NCII and DCII) and HLA in rheumatoid arthritis (RA). METHODS: One hundred fourteen patients with clinically well-defined RA were HLA-DR and DQ typed. Those who were DR4 positive were subtyped for DRB1*0401-*0408 alleles by polymerase chain reaction using allele-specific oligonucleotide probes. Antibodies to human NCII and DCII (heat-denatured) were measured by enzyme-linked immunosorbent assay. The frequency of HLA alleles was compared in patients grouped according to the presence and absence of antibodies to NCII and DCII. RESULTS: Twenty-seven patients (24%) were positive for antibodies to NCII. There was a significant increase in the frequency of HLA-DR7 in anti-NCII-positive patients compared with anti-NCII-negative patients (30% versus 9%; P = 0.019) and a significant decrease in HLA-DR3 (7% versus 28%; P = 0.044). Repeating the analyses after excluding the 16 patients who were DR7 positive revealed a significant increase in the frequency of HLA-DR1 in anti-NCII-positive patients compared with anti-NCII-negative patients (63% versus 27%; P = 0.045). Moreover, antibodies to NCII were associated with the third hypervariability region susceptibility sequence QRRAA that is present in DRB1*0101, *0404, *0405, and *0408 (84% versus 47%; P = 0.0085); 24 of 27 anti-NCII-positive patients were positive for either DR7, DR1, or DRB1*0404 or *0408. Thirty patients (26%) were positive for antibodies to DCII. There was a significant increase in the frequency of HLA-DR3 in anti-DCII-positive patients compared with anti-DCII-negative patients (40% versus 18%; P = 0.028). CONCLUSION: The genetic associations between HLA-DR alleles and antibodies to CII in RA patients is in keeping with the collagen-induced arthritis model and implicates autoimmunity to CII as a major component in the multifactorial pathogenesis of RA.

Mimotopes identified by phage display for the monoclonal antibody CII-C1 to type II collagen.

The characterization of B cell epitopes has been advanced by the use of random peptide libraries displayed within the coat protein of bacteriophage. This technique was applied to the monoclonal antibody (mAb) C1 to type II collagen (CII-C1). CII-C1 is known to react with a conformational epitope on type II collagen that includes residues 359-363. Three rounds of selection were used to screen two random nonameric phage libraries and 18 phagotopes were isolated. CII-C1 reacted by ELISA with 17 of the 18 phagotopes: one phagotope contained a stop codon. Of the eight most reactive phage, seven inhibited the reactivity by ELISA of CII-C1 with type II collagen. Of the 18 phage isolated, 11 encoded the motif F-G-x-Q with the sequence F-G-S-Q in 6, 2 encoded F-G-Q, and one the reverse motif Q-x-y-F. Most phagotopes that inhibited the reactivity of CII-C1 encoded two particular motifs consisting of two basic amino acid residues and a hydrophobic residue in the first part of the insert and the F-G-x-Q or F-G-Q motif in the second part; phagotopes which contained only one basic residue in the first part of the sequence were less reactive. These motifs are not represented in the linear sequence of type II collagen and thus represent mimotopes of the epitope for CII-C1 on type II collagen. There were five phagotopes with peptide inserts containing the sequence RLPFG occurring in the Epstein-Barr virus nuclear antigen, EBNA-1. This is of interest because EBV has been implicated in the initiation of rheumatoid arthritis (RA) by reason of increased reactivity to EBNA-1 in RA sera. In conclusion, the phage display technique disclosed mimotopes for a conformational epitope of type II collagen, and revealed an interesting homology with a sequence of the EBNA-1 antigen from Epstein Barr virus.

Psychiatric treatment of children and adolescents in rural communities. Myths and realities.

Rural child and adolescent psychiatry offers many challenges, a varied and interesting practice, and the satisfaction of performing needed and important work in an environment in which one's presence is valued. The successful psychiatrist can expect to be an integrated and appreciated member of the community. The fit is not a good one for every practitioner, however. Not only are incomes lower, although the cost of living is low as well, but practitioners may find they have only exchanged urban stresses for rural pressures. The characteristics important for the child and adolescent psychiatrist are the same for rural and urban settings: flexibility, creativity and innovation, competence, self confidence, a good sense of boundaries, a good balance between personal and private life, supportive personal relationships, and a sense of humor. One must be a child advocate, have a willingness to give of one's self and one's time, and be down to earth, comfortable with oneself, and capable of self entertainment. Training programs with access to rural populations can introduce residents to rural child and adolescent psychiatry while supporting those who are already in practice. The authors hope that this article will promote a dialogue with psychiatrists considering relocation to a rural area and encourage training programs to prepare residents for rural practice.

Characterization and development of RGD-peptide-modified poly(lactic acid-co-lysine) as an interactive, resorbable biomaterial.

The design of biomaterials containing specific ligands on the surface offers the possibility of creating materials that can interact with and potentially control mammalian cell behavior. Biodegradable materials further provide the significant advantage that the polymer will disappear in vivo, obviating long-term negative tissue responses as well as the need for retrieval. In earlier studies we synthesized and characterized arginine-glycine-aspartic acid (RGD) peptide-modified poly(lactic acid-co-lysine) (PLAL). In this study, both bulk properties and surface features have been characterized, with a focus on surface analysis as a means of interpreting observed changes in cell behavior. Bulk peptide attachments were performed using 1,1'-carbonyldiimidazole (CDI). Amino groups were measured using colorimetric assays and X-ray photoelectron spectroscopy (XPS). Peptides were measured by incorporating iodine into the peptide as a distinct elemental marker for use with XPS. Typical samples contained 13 +/- 4 pmol/cm2 of amino groups and 4 +/- 0.2 pmol/ cm2 of peptides, as calculated from XPS measurements of nitrogen and iodine. The wettability and crystallinity of the samples were determined by contact angles and differential scanning calorimetry, respectively. Wettability and crystallinity were not altered by the incorporation of lysine or peptides. After incubating bovine aortic endothelial (BAE) cells for 4 h on surfaces with RGD-containing peptides, the mean spread cell area increased from 77 +/- 2 microns2 to 405 +/- 29 microns2 compared to 116 +/- 11 microns2 on poly(lactic acid), 87 +/- 4 microns2 on PLAL, and 105 +/- 4 microns2 on surfaces with RDG-containing (control) peptides. The significance of this work is that the first synthetic interactive, resorbable biomaterial has been developed, and use of this material to control cell behavior has been demonstrated.

Measurement of exposure to mouse urinary proteins in an epidemiological study.

OBJECTIVES: To develop an assay to measure airborne mouse urinary protein (MUP) and to assess the occupational exposure to MUP in the workforce of three establishments as part of an epidemiological study examining the influence of aeroallergen exposure on the development of allergic respiratory disease. METHODS: Personal air samples were collected from nine exposure groups during a workshift. A sensitive and reproducible competitive inhibition assay, which used rabbit antisera specific for MUP, was developed and used to measure the occupational exposure to MUP. RESULTS: The personal measurements of MUP showed that people with direct contact with mice (animal technicians) had the highest exposure followed in decreasing order by those working with anaesthetised animals or their tissue (postmortem workers and scientists) and those with indirect contact with mice (supervisors, office workers, and slide production workers). The only difference in concentrations of MUP between the three establishments were found for cage cleaners, which reflected differences in working practises for this exposure category. Air samples collected during the performance of specific tasks showed that high exposures to MUP were associated with handling mice, indirect contact with mice, and washing floors. CONCLUSIONS: Exposure to mouse urinary proteins has been measured in the occupational environment. This information can be used to determine the relation between exposure to MUP and the development of allergic and respiratory disease.

Colorimetric analysis of surface reactive amino groups on poly(lactic acid-co-lysine):poly(lactic acid) blends.

The quantification of functional amino (NH2) groups on poly(lactic acid-co-lysine):(poly(L-lactic acid (PLAL:PLA) blends was performed using a colorimetric assay based on the reaction of sulpho-succinimidyl-4-O-(4,4'-dimethoxytrityl)-butyrate (sulpho-SDTB) with primary amino groups. The colorimetric assay was used to assess the available reactive sites for coupling of biologically active species to PLAL. Blends were created that contained from 10 to 70 wt% poly(lactic acid-co-lysine). Bulk lysine contents within the blends were determined by amino acid analysis and ranged from 9.1 micromol g(-1) to 52.9 micromol g(-1) for blends created using PLA of 100000g mol(-1) molecular weight. Surface amino group concentrations on the same set of blends ranged from 0.23 to 1.45 nmol cm(-2). Similar surface amino groups concentrations were measured on blends using 50000, 200000 and 300000g mol(-1) poly(lactic acid). Non-specific interactions of the colorimetric assay reagents with the PLAL-containing blends were measured on blends prepared from epsilon-amino protected PLAL and 100000g mol(-1) PLA. The presence of amino groups within the top 50 angstroms was confirmed by X-ray photoelectron spectroscopy.

IgG subclasses of antibodies to type II collagen in rheumatoid arthritis differ from those in systemic lupus erythematosus and other connective tissue diseases.

OBJECTIVE: To compare IgG subclass distribution of autoantibodies to native type II collagen in patients with rheumatoid arthritis (RA) and in other rheumatic and inflammatory diseases including systemic lupus erythematosus (SLE). METHODS: The IgG subclass of antibodies to native type II collagen were measured by modified ELISA using chemiluminescent detection and subclass-specific monoclonal antibodies in sera that contained IgG antibodies to collagen using conventional ELISA. RESULTS: Antibodies to native type II collagen were present in sera of 20% of 323 patients with RA. 9% of 211 patients with SLE, 12% of 50 patients with osteoarthritis, and 13% of 75 patients with other chronic inflammatory diseases, but the highest levels occurred in patients with RA. The IgG subclass distribution of these antibodies differed markedly according to disease. In RA the predominant subclasses were IgG1 (58%) and IgG3 (47%), whereas in SLE the predominant subclass was IgG4 (69%). IgG2 was represented across all disease groups. Overall the frequency of IgG1 and/or IgG3 antibodies to type II collagen (84%) was significantly greater in RA than in other disease groups combined (20%) (p < 0.0001). CONCLUSION: Antibodies to native collagen in RA were predominantly of the complement fixing and potentially damaging subclasses IgG1 and IgG3, but in SLE were of the non-complement fixing subclass IgG4; in other diseases, where present, antibodies were mostly IgG2 and of low level. These observations support the importance of autoimmunity to collagen in the pathogenesis of RA.

Antibodies to type II collagen in early rheumatoid arthritis. Correlation with disease progression.

OBJECTIVE: To establish that frequencies and levels of IgG antibodies to type II collagen are higher in early rheumatoid arthritis (RA), and to correlate these results with disease activity. METHODS: Forty-four patients were characterized as having early RA. Patient sera obtained at initial presentation and at 12 months were tested by enzyme-linked immunosorbent assay for IgG antibodies to native and denatured type II collagen. RESULTS: IgG antibodies to native and denatured type II collagen were detected at initial presentation in 27% and 82% of patients, respectively, and after 12 months in 14% and 50%, respectively. The presence of antibodies to native collagen was associated with activity of RA and severity of symptoms, and loss of antibodies at 12 months was associated with initially erosive RA and the DRB1 disease susceptibility motif. CONCLUSION: Levels of serum IgG antibodies to collagen in RA decrease over time and, therefore, are not attributable simply to cartilage destruction. The presence of early positivity for these antibodies, together with the RA susceptibility motif, appears to be predictive of rapidly progressive RA.

A major outbreak of asthma associated with a thunderstorm: experience of accident and emergency departments and patients' characteristics. Thames Regions Accident and Emergency Trainees Association.

OBJECTIVE: To investigate the time course of an epidemic of asthma after a thunderstorm, characteristics of patients affected, and the demand on emergency medical resources. DESIGN: Study of registers and records in accident and emergency departments and questionnaire to staff. SETTING: London area. SUBJECTS: All patients presenting at 12 accident and emergency departments with asthma or other airway disease. MAIN OUTCOME MEASURES: Numbers of patients, clinical features, information on shortage of resources--equipment, drugs and staff. RESULTS: The epidemic had a sudden onset on 24 June 1994; 640 patients with asthma or other airways disease attended during 30 hours from 1800 on 24 June, nearly 10 times the expected number. Over half (365) the patients were aged 21 to 40 years. A history of hay fever was recorded in 403 patients; for 283 patients this was the first known attack of asthma; a history of chronic obstructive airways disease was recorded in 12 patients. In all, 104 patients were admitted (including five to an intensive care unit). Several departments ran out of equipment or drugs, called in additional doctors, or both. CONCLUSIONS: This study supports the view that this epidemic was larger than previously reported epidemics and the hypothesis that "thunderstorm associated asthma' is related to aeroallergens. Demands on resources were considerable; a larger proportion of patients needing intensive care would have caused greater problems.

Comparative epitope mapping of antibodies to collagen in women with silicone breast implants, systemic lupus erythematosus and rheumatoid arthritis.

Previous work has shown that women with silicone gel breast implants have an increased frequency of autoantibodies to collagen types I and II. 70 women without a specific autoimmune disease, using criteria of the American College of Rheumatology, but who had silicone breast implants were studied for the presence of serum antibodies to native and denatured human types I and II collagen by ELISA. 82 women with systemic lupus erythematosus (SLE), 94 women with rheumatoid arthritis (RA), and 133 healthy controls were also studied. There was a high frequency of autoantibodies to collagen in each of the groups when compared to the healthy controls. The specificities of these antibodies were found to differ markedly when examined by immunoblotting using peptides derived by cyanogen bromide digestion of the collagens. Sera from women with silicone implants reacted with multiple peptides of type I collagen in an individual-specific manner, but sera from women with SLE or RA reacted weakly with a restricted range of peptides. Against type II collagen, sera from women with RA reacted strongly with multiple peptides, while sera from women with silicone implants or SLE reacted only weakly or not at all. The patterns of reactivity against collagens by sera from women with silicone implants suggest that silicone can act as an adjuvant to enhance the immunogenicity of type I collagen.

Antibodies to collagen: comparative epitope mapping in women with silicone breast implants, systemic lupus erythematosus and rheumatoid arthritis.

Women with silicone breast implants have a significantly increased frequency of antibodies to collagen types I and II. To characterize the specificity of these antibodies, 70 women without a specific autoimmune disease, according to the criteria of the American College of Rheumatology, but who had silicone breast implants were studied for the presence of serum antibodies to native and denatured human types I and II collagen by ELISA. Positive sera were further studied by immunoblotting using peptides derived by cyanogen bromide digestion of the collagens. Samples of 82 women with systemic lupus erythematosus (SLE), 94 women with rheumatoid arthritis (RA), and 133 healthy controls were studied concurrently. There was a high frequency of autoantibodies to collagen in each of the study groups when compared to the healthy controls. However, and of particular interest, the epitope specificity of the autoantibodies differed markedly. Sera from women with silicone implants reacted strongly in an individual-specific manner with multiple peptides of type I collagen, whereas sera from women with SLE and RA reacted only weekly with a restricted range of peptides of type I collagen. Sera from women with RA reacted strongly with multiple peptides of type II, whereas sera from women with silicone implants or SLE reacted only weakly. The reactivity of women with silicone implants suggests that silicone or its biodegradation products can act as adjuvants in situ to enhance the immunogenicity of type I collagen, or protein-silicone conjugates.

Porphobilinogen deaminase is encoded by a single gene in Arabidopsis thaliana and is targeted to the chloroplasts.

Porphobilinogen deaminase (PBG deaminase) is an early enzyme of the pathway for chlorophyll and heme synthesis. Using degenerate oligonucleotide primers, based on amino acid sequence data for purified PBG deaminase from pea, a fragment was amplified from Arabidopsis genomic DNA by PCR, and then used to isolate both a cDNA and a genomic clone for PBG deaminase from Arabidopsis. The cDNA, shown to be full-length by primer extension, encodes a precursor protein of 382 residues, which can be imported into isolated chloroplasts and processed to the mature size. The genomic clone encodes an identical sequence to the cDNA, except for the presence of four introns within the coding region of the mature protein, and 1.7 kb of upstream sequence. There is no obvious TATA box within 50 bp of the transcription start. Southern blot analysis suggests that PBG deaminase is encoded by a single gene in the Arabidopsis genome, and RNase protection experiments demonstrated that this gene is expressed in both leaves and roots. These results support the conclusion that there is only one form of PBG deaminase in all plant cells, which is located in the plastid.

Specificity of antibodies to type II collagen in early rheumatoid arthritis.

OBJECTIVE: To analyze the antibody response to native type II collagen in early rheumatoid arthritis (RA), examining the immunoglobulin isotypes, and polypeptide epitopes recognized, in patients followed over a 2-year period from within 6 months of the first occurrence of symptoms. METHODS: Sera from 16 patients were studied, of whom 10 had antibodies to native type II collagen and 6 did not. The clinical and laboratory assessment, carried out initially and at 6 monthly intervals included the number of 1958 ARA criteria fulfilled, Ritchie index, erythrocyte sedimentation rate, rheumatoid factor and radiological assessment. An ELISA was used to measure IgG, IgA and IgM antibodies, and immunoblotting to identify the number and location of epitopes, using polypeptides prepared by cyanogen bromide digestion of human type II collagen. RESULTS: Antibodies to type II collagen were present in all sequential serum samples for the 10 antibody positive patients. None of the 6 patients who initially lacked antibodies developed them. The antibodies were of IgG isotype in 9, of IgA isotype in 8, and of IgM isotype exclusively in one. At the initial clinical assessment patients with antibodies to collagen were indistinguishable from those without. At 12 and 24 months patients with antibodies fulfilled significantly more ARA criteria than antibody negative patients. The patterns of antibody reactivity to collagen polypeptides by immunoblotting were constant over time but differed from patient to patient. CONCLUSION: The presence of an established and persisting IgG antibody response to type II collagen in early RA before cartilage destruction is evident points to a subset of RA, perhaps equivalent to the collagen induced model in animals, in which this immune response is intrinsic to pathogenesis.

Antibodies to the collagen-like region of C1q and type II collagen are independent non-cross-reactive populations in systemic lupus erythematosus and rheumatoid arthritis.

The collagen-like region (CLR) of the first component of complement, C1q, and type II collagen are structurally similar, raising the possibility of epitopes in common, and of the existence of autoantibodies that are cross-reactive. Accordingly, antibodies to the CLR of C1q and to type II collagen were measured in patients' sera with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) by an ELISA. IgG antibodies to the CLR of C1q were present in 17% of patients with SLE but none with RA, and IgA antibodies were present in 10% and 8% of patients with SLE and RA, respectively. IgG antibodies to type II collagen were present in 15% and 25% of patients with SLE and RA, respectively, and IgA antibodies in 15% and 28% of patients with SLE and RA, respectively. There was no correlation in either disease between the serum levels of antibodies to the CLR of C1q and antibodies to type II collagen. For sera with antibodies to both antigens, neither competitive inhibition by ELISA nor preabsorption with the alternative antigen affected the level of reactivity to the other antigen. Thus antibodies to the CLR of C1q and antibodies to type II collagen are independent and non-cross-reactive populations, and presumably occur by different types of immunogenic stimulation.