169 results on '"Braspenning, J"'
Search Results
152. Blood pressure control in treated hypertensive patients: clinical performance of general practitioners.
- Author
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Frijling BD, Spies TH, Lobo CM, Hulscher ME, van Drenth BB, Braspenning JC, Prins A, van der Wouden JC, and Grol RP
- Subjects
- Adolescent, Adult, Aged, Aged, 80 and over, Cross-Sectional Studies, Female, Guideline Adherence, Humans, Hypertension physiopathology, Male, Middle Aged, Netherlands, Practice Guidelines as Topic, Task Performance and Analysis, Antihypertensive Agents therapeutic use, Blood Pressure drug effects, Clinical Competence, Family Practice standards, Hypertension drug therapy
- Abstract
Background: The blood pressure of many treated hypertensive patients remains above recommended target levels. This discrepancy may be related to general practitioners' (GPs') actions., Aim: To assess clinical performance of GPs in blood pressure control in treated hypertensive patients and to explore the influence of patient and GP characteristics on clinical performance., Design of Study: Cross-sectional study conducted on 195 GPs with invitations to participate made via bulletins and by letter., Setting: One hundred and thirty-two practices in the southern half of The Netherlands from November 1996 to April 1997., Method: Performance criteria were selected from Dutch national hypertension guidelines for general practice. GPs completed self-report forms immediately after follow-up visits of hypertensive patients treated with antihypertensive medication., Results: The GPs recorded 3526 follow-up visits. In 63% of these consultations the diastolic blood pressure (DBP) was 90 mmHg or above. The median performance rates of the GPs were less than 51% for most of the recommended actions, even at a DBP of > or = 100 mmHg. Performance of non-pharmacological actions increased gradually with increasing DBP; prescribing an increase in antihypertensive medication and making a follow-up appointment scheduled within six weeks rose steeply at a DBP of > or = 100 mmHg. Patient and GP characteristics contributed little to clinical performance. Action performance rates varied considerably between GPs., Conclusion: GPs seem to target their actions at a DBP of below 100 mmHg, whereas guidelines recommend targeting at a DBP of below 90 mmHg.
- Published
- 2001
153. Low back pain in general practice: reported management and reasons for not adhering to the guidelines in The Netherlands.
- Author
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Schers H, Braspenning J, Drijver R, Wensing M, and Grol R
- Subjects
- Adolescent, Adult, Age Factors, Family Practice, Female, Humans, Male, Middle Aged, Netherlands epidemiology, Prospective Studies, Treatment Outcome, Low Back Pain diagnosis, Low Back Pain therapy, Practice Guidelines as Topic
- Abstract
Background: Although guidelines for the management of low back pain have been published in the past decade, there is potential for further improvement in back pain care., Aim: To document the management of non-specific low back pain by general practitioners (GPs) in the Netherlands, to determine how this management of care is related to patient and physician factors, and to explore possible reasons for not adhering to the guidelines., Method: A prospective study was set up in which 57 GPs in 30 general practices completed a computerised questionnaire after each consultation for low back pain during a four-month period., Results: Of 1640 back pain contacts, 1180 referred to non-specific low back pain. Diagnostic tests were ordered in 2% of first consultations and in 7% of follow-up consultations within one episode. The advice to stay active despite pain was given in 76% and 69% of these cases respectively. Patients were prescribed an analgesic in 53% and 41% of cases respectively (mainly NSAIDs [80%]). Patients were referred to a physiotherapist in 22% of first and in 50% of follow-up consultations. Older patients were physically examined less often, prescribed analgesics more often, and were told less often that staying active could benefit them. The advice to remain active was omitted more often when symptoms lasted longer. Only a small part of the variance in management was accounted for by patient characteristics or by differences between practices., Conclusion: The management of low back pain met the guidelines to a large extent. Management decisions were often related to characteristics in which the guidelines lack differentiation. Important reasons for non-adherence were perceived patients' preferences. Further implementation of guidelines will be difficult unless doctors' and patients' views are more explicitly known.
- Published
- 2000
154. Perceived barriers to the implementation of diabetes guidelines in hospitals in The Netherlands.
- Author
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Dijkstra RF, Braspenning JC, Uiters E, van Ballegooie E, and Grol RT
- Subjects
- Attitude of Health Personnel, Diabetes Mellitus epidemiology, Guideline Adherence, Humans, Netherlands epidemiology, Practice Guidelines as Topic, Surveys and Questionnaires, Diabetes Mellitus therapy
- Abstract
Objective: To determine the organisational and personal barriers to the implementation of diabetes guidelines in hospitals in The Netherlands and relate them to structural factors of diabetes care., Method: In a written survey internists specialised (or with a specific interest) in diabetes in all general hospitals in The Netherlands (n = 120) were asked to indicate the perceived organisational and personal barriers to adherence to the diabetes guidelines. In the same questionnaire their activities related to diabetes care and the working hours of the additional personnel involved were measured., Results: There was at least one specialised diabetes nurse employed in all hospitals, although the extent of the appointment varied widely from 0.2 to 6.9 full-time equivalent (average 1.5). In most hospitals (90%) a diabetes care team had been established, while podiatrists were working in only 72% of the hospitals. Furthermore, 65-80% of the hospitals organised special consultation hours for diabetic patients, had a protocol for diabetes treatment, or patient held administration booklets. The most frequently mentioned barriers to the implementation of diabetes guidelines were high workload, no adequate financial compensation, and a shortage of necessary personnel., Conclusion: A number of preconditions for structured diabetes care, like the presence of a diabetes team and a specialised diabetes nurse, were in place. However, large differences between the hospitals in the organisation of diabetes care and the availability of staff, together with the related perceived barriers to the implementation of the guidelines showed that there are still many opportunities for improvements.
- Published
- 2000
- Full Text
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155. Attendance to cervical cancer screening in family practices in The Netherlands.
- Author
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Hermens RP, Tacken MA, Hulscher ME, Braspenning JC, and Grol RP
- Subjects
- Adult, Community Health Centers statistics & numerical data, Cross-Sectional Studies, Female, Humans, Middle Aged, Netherlands epidemiology, Uterine Cervical Neoplasms epidemiology, Family Practice statistics & numerical data, Mass Screening methods, Patient Compliance statistics & numerical data, Uterine Cervical Neoplasms prevention & control
- Abstract
Background: The effectiveness of three different organizational approaches to cervical cancer screening (community based, family practice based, and a combination) was evaluated in nationally representative family practices., Method: We selected 122 family practices with a computerized sex-age register from a database of 1, 251 family practices, representative of all 4,758 family practices in The Netherlands. Approximately 40 practices were linked with each approach. We measured the attendance, the reasons for nonattendance, and the influence of a reminder on the attendance of women invited for cervical screening in September, October, and November 1996. The patients were grouped according to age. A cross-sectional design was used for the study., Results: For younger women, the total attendance rate, coverage (percentage of women "protected" against cervical cancer), and control rate (percentage of women with medical reasons for nonattendance or postponement of the smear) were highest in practices using the family practice-based approach (68, 77, and 90%, respectively) and lowest in practices with the community-based approach (53, 62, and 68%, respectively). For older women, the family practice-based approach and the combination approach were associated with attendance rates significantly higher than those for the community-based approach (approximately 60, 80, and 80% vs 47, 67, and 70%, respectively). A reminder sent by the family physician to women not responding to an initial invitation increased the attendance rate by 7 to 11% in both age categories, depending on who had sent the first invitation., Conclusion: A family practice-based cervical screening approach appeared to be the most effective at a national level, achieving the highest attendance rate, coverage, and control rate., (Copyright 2000 American Health Foundation and Academic Press.)
- Published
- 2000
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156. Improving population-based cervical cancer screening in general practice: effects of a national strategy.
- Author
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Hermens RP, Hak E, Hulscher ME, Mulder J, Tacken MA, Braspenning JC, and Grol RP
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- Documentation, Female, Humans, Logistic Models, Netherlands, Odds Ratio, Program Evaluation, Prospective Studies, Registries, Reminder Systems, Family Practice standards, Guideline Adherence, Mass Screening organization & administration, National Health Programs organization & administration, Uterine Cervical Neoplasms prevention & control
- Abstract
Objective: To assess the effects of a Dutch national prevention programme, aimed at general practitioners (GPs), on the adherence to organizational guidelines for effective cervical cancer screening in general practice. To identify the characteristics of general practices determining success., Design: A prospective questionnaire study with pre- and post-measurement (before and 15 months after the introduction of the national programme)., Setting and Study Participants: A random sample of one-third of all 4758 Dutch general practices. One GP was asked to participate per practice., Intervention: A national GP prevention programme to improve population-based prevention of cervical cancer combining various methods for quality improvement in general practice, performed on a national, district and practice level. Outreach visitors were a key strategy in bringing about behavioural changes., Main Outcome Measures: The proportion of practices adhering to 10 recommendations (in four guidelines) to organize effective cervical cancer screening., Results: After 15 months, all Dutch practices showed significant improvement in adherence to nine out of 10 recommendations. Two recommendations, in particular 'identifying women who should be medically excluded from screening' and 'sending a reminder to non-compliers' showed the largest absolute increases of 26% and 33%, respectively. Besides more intensive support of outreach visitors, practice characteristics such as 'computerization' and 'delegation of many clinical tasks to the practice assistant' were important in improving the adherence to guidelines., Conclusion: The national programme, with a combination of various methods for quality improvement, appeared to be effective in improving the organization of cervical screening in general practice. Computerization and, to a lesser extent, delegation of many clinical tasks to the practice assistant and more intensive support to practices, positively influenced the effectiveness of the national programme.
- Published
- 1999
- Full Text
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157. Dutch GPs acknowledge the need for preconceptual health care.
- Author
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Cikot R, Gaytant M, Steegers E, and Braspenning J
- Subjects
- Female, Humans, Netherlands, Family Practice education, Health Promotion organization & administration, Preconception Care methods
- Published
- 1999
158. E7-specific cytotoxic T cell tolerance in HPV-transgenic mice.
- Author
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Borchers A, Braspenning J, Meijer J, Osen W, Gissmann L, and Jochmus I
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- Animals, Antibodies, Viral biosynthesis, Immunization, Mice, Mice, Inbred C3H, Mice, Inbred C57BL, Mice, Transgenic, Oncogene Proteins, Viral genetics, Papillomavirus E7 Proteins, Transgenes, Immune Tolerance, Oncogene Proteins, Viral immunology, Papillomaviridae immunology, Repressor Proteins, T-Lymphocytes, Cytotoxic immunology
- Abstract
The "high-risk" human papillomavirus type 16 (HPV 16) is associated with the development of cervical cancer. Although the viral gene products E6 and E7 are constitutively expressed in HPV 16-associated lesions and therefore appear as candidate antigens for a specific immune response, the immune system fails to produce an efficient defence against tumor outgrowth in affected patients. Keratinocytes are the natural target cells of HPV infection. To investigate the E7-specific immune response in vivo, we used transgenic mice expressing the oncogenes E6 and E7 of HPV 16 under the control of the keratin 10 promoter in the suprabasal layers of the epidermis. This expression pattern closely reflects the viral early gene transcription that is observed in low grade cervical intraepithelial lesions (CIN). The transgene product E7 does not induce an immune response in these transgenic mice. However, upon vaccination anti-E7 antibodies were produced without causing signs of autoimmune disease. In contrast, E7-specific cytotoxic T lymphocytes (CTL) were not detected after immunization. From these results we conclude that in K10 HPV 16 E6/E7 transgenic mice the E7 transgene expression induces specific immunological tolerance on the CTL level.
- Published
- 1999
- Full Text
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159. [Preconception counseling in family practice; a survey of 100 family physicians].
- Author
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Gaytant MA, Cikot RJ, Braspenning JC, Grol RP, Merkus JM, and Steegers EA
- Subjects
- Data Collection, Female, Health Knowledge, Attitudes, Practice, Humans, Male, Netherlands, Population Surveillance, Pregnancy, Preventive Health Services organization & administration, Family Planning Services education, Family Practice statistics & numerical data, Prenatal Care statistics & numerical data
- Abstract
Objective: To assess the awareness, opinions on desirability and preconditions such as special knowledge with regard to the concept of preconceptional health counselling among general practitioners., Design: Descriptive., Setting: General practices in the Groot Gelre district of the Dutch General Practitioners Society., Method: A representative sample of 100 general practitioners were asked by phone for permission to send a questionnaire; 89 out of the 94 general practitioners who were sent a questionnaire replied (response: 89%)., Results: Almost all general practitioners (88%) knew about the concept of preconceptional health counselling and most of them already gave some kind of preconceptional advice. 93% considered preconceptional health counselling part of their job responsibility and 91% were prepared to give more preconceptional health care in the future. 53% of all general practitioners, however, indicated that they lacked sufficient knowledge to give adequate advice. The advantages of preconceptional health care were considered to outweigh possible disadvantages such as medicalization of pregnancy., Conclusion: The general practitioners knew about preconceptional health care and considered it part of their job. Many already provided preconceptional health care, although not in a structured way. General practitioners appeared to lack time and appropriate knowledge, which indicates a need for postgraduate training.
- Published
- 1998
160. Secretion of heterologous proteins from Schizosaccharomyces pombe using the homologous leader sequence of pho1+ acid phosphatase.
- Author
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Braspenning J, Meschede W, Marchini A, Müller M, Gissmann L, and Tommasino M
- Subjects
- Fluorescence, Green Fluorescent Proteins, Luminescent Proteins genetics, Luminescent Proteins metabolism, Microscopy, Fluorescence, Oncogene Proteins, Viral metabolism, Papillomavirus E7 Proteins, Phosphoproteins metabolism, Protein Sorting Signals genetics, Recombinant Fusion Proteins isolation & purification, Recombinant Fusion Proteins metabolism, Schizosaccharomyces enzymology, Viral Proteins metabolism, Acid Phosphatase chemistry, Fungal Proteins metabolism, Protein Sorting Signals metabolism, Schizosaccharomyces metabolism
- Abstract
In this study we report the use of the S. pombe leader sequence of pho1+ acid phosphatase (Elliott et al., J. Biol. Chem. 216, 2916-2941, 1986) for the secretion of heterologous proteins into the medium. The green fluorescent protein (GFP) and the Human Papillomavirus (HPV) type 16 E7 protein are normally not secreted; fusion of the S. pombe pho1 leader peptide (SPL) to GFP and HPV 16 E7 resulted in an efficient secretion of these proteins although the latter contains a nuclear targeting sequence. These data suggest that SPL fused constructs could be applied for the production of other recombinant proteins using the S. pombe expression system. Furthermore, since GFP retains its intrinsic fluorescence during the secretion, this system may be useful to study the secretory pathway of fission yeast in vivo., (Copyright 1998 Academic Press.)
- Published
- 1998
- Full Text
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161. Do general practices adhere to organizational guidelines for effective cervical cancer screening?
- Author
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Hermens RP, Hak E, Hulscher ME, Mulder J, Braspenning JC, and Grol RP
- Subjects
- Community Health Services, Data Collection, Female, Humans, Netherlands, Random Allocation, Guideline Adherence statistics & numerical data, Physicians, Family statistics & numerical data, Practice Guidelines as Topic, Uterine Cervical Neoplasms diagnosis
- Abstract
Background: Well-organized cervical screening has been shown to be effective in the reduction of both morbidity and mortality from cancer of the uterine cervix. In The Netherlands, the GP plays an important role in the cervical screening. The question is whether the general practices are able to organize an effective cervical cancer screening., Objectives: We explored the extent to which Dutch general practices adhere to organizational guidelines for effective population-based prevention of cervical cancer and which practice characteristics are important for this adherence., Methods: A postal survey was conducted in a random sample of one-third of all 4758 Dutch general practices. Two sets of information were collected: practice characteristics and adherence to four organizational guidelines for effective cervical screening concerning inviting the women, monitoring attendance and sending reminders, organizing the taking of the smear and follow-up monitoring., Results: A total of 1251 (79%) general practices returned a questionnaire; 90 questionnaires were excluded from analyses owing to missing data. The 1161 practices were representative of the Netherlands. A minority of the practices adhered to the four guidelines (in total, ten recommendations). The presence of the practice characteristics 'a general practice-based inviting system', 'a high delegation index' (delegating many tasks to the assistants) and a 'computerized patient information recording system' were positively associated with the adherence to most of the guidelines., Conclusion: This study showed that most of the Dutch general practices are not yet ready to organize an effective cervical cancer screening system. A general practice-based inviting system, a high delegation index and a computerized patient information recording system proved to be important for the adherence to the guidelines. In order to organize a cervical screening programme to achieve optimal effectiveness, emphasis should be placed on the adherence to the four guidelines described in this study and on stimulating a general practice-based inviting system, delegation to the practice assistant and computerization.
- Published
- 1998
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162. The CXXC Zn binding motifs of the human papillomavirus type 16 E7 oncoprotein are not required for its in vitro transforming activity in rodent cells.
- Author
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Braspenning J, Marchini A, Albarani V, Levy L, Ciccolini F, Cremonesi C, Ralston R, Gissmann L, and Tommasino M
- Subjects
- 3T3 Cells metabolism, Adenovirus E1A Proteins metabolism, Adenovirus E1A Proteins physiology, Amino Acid Sequence, Animals, Binding Sites, Conserved Sequence, Dimerization, Mice, Oncogene Proteins, Viral physiology, Papillomaviridae, Papillomavirus E7 Proteins, Retinoblastoma Protein metabolism, Retinoblastoma Protein physiology, Zinc Fingers, Cell Transformation, Viral physiology, Oncogene Proteins, Viral metabolism, Zinc metabolism
- Abstract
The conserved region 3 (CR3) of the E7 protein of human papillomaviruses contains two CXXC motifs involved in zinc binding and in the homodimerization of the molecule. Studies have suggested that the intact CXXC motifs in the CR3 of HPV16 and HPV18 E7 are required for the in vitro transforming activity of these proteins. CR3 also contains a low affinity pRb binding site and is involved in the disruption of the E2F/Rb1 complex. E7 is structurally and functionally related to Adenovirus E1A protein, which also has two CXXC motifs in CR3. However, the Ad E1A transforming activity appears to be independent of the presence of such domains. In fact, this viral protein exists in vivo as two different forms of 289 and 243 amino acids. The shorter Ad E1A form (Ad E1A243), where both CXXC motifs are deleted by internal splicing, retains its in vitro transforming activity. We have investigated if the HPV16 E7 CR3 can be functionally replaced by the Ad E1A243 CR3, which lacks both CXXC motifs. A chimeric protein (E7/E1A243) containing the CR1 and CR2 of HPV16 E7 fused to the CR3 of Ad E1A 243 was constructed. The E7/E1A243 while not able to homodimerize in the S. cerevisiae two-hybrid system retains several of the properties of the parental proteins, HPV16 E7 and Ad E1A. It associates with the 'pocket' proteins, induces growth in soft agar of NIH3T3 cells and immortalizes rat embryo fibroblasts. These data suggest that the CXXC motifs in CR3 of E7 do not play a direct role in the transforming properties of this viral protein but probably are important for maintaining the correct protein configuration.
- Published
- 1998
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163. Antibodies against early proteins of human papillomaviruses as diagnostic markers for invasive cervical cancer.
- Author
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Meschede W, Zumbach K, Braspenning J, Scheffner M, Benitez-Bribiesca L, Luande J, Gissmann L, and Pawlita M
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- Adult, Aged, Antigens, Viral genetics, Antigens, Viral immunology, Female, Humans, Middle Aged, Oncogene Proteins, Viral genetics, Oncogene Proteins, Viral immunology, Papillomaviridae genetics, Papillomavirus E7 Proteins, Papillomavirus Infections blood, Papillomavirus Infections immunology, Recombinant Proteins immunology, Sensitivity and Specificity, Tumor Virus Infections blood, Tumor Virus Infections immunology, Uterine Cervical Neoplasms blood, Antibodies, Viral immunology, DNA-Binding Proteins, Enzyme-Linked Immunosorbent Assay methods, Papillomaviridae immunology, Papillomavirus Infections diagnosis, Repressor Proteins, Tumor Virus Infections diagnosis, Uterine Cervical Neoplasms virology
- Abstract
Cervical cancer is the most prevalent tumor in developing countries and the second most frequent cancer among females worldwide. Specific human papillomaviruses (HPVs) and, most notably, HPV types 16 and 18 are recognized as being causally associated with this malignancy. Antibodies against early HPV proteins E6 and E7 have been found more often in patients with tumors than in controls. Existing peptide enzyme-linked immunosorbent assays (ELISAs) for the detection of anti-E6 and anti-E7 antibodies in human sera have low levels of sensitivity and specificity and thus are not suitable for use as diagnostic tools. Based on highly purified recombinant native proteins, we developed four sandwich ELISAs for the detection of antibodies against HPV type 16 and 18 E6 and E7 proteins. We demonstrate their sensitivities and high degrees of specificity for cervical cancer. Among a total of 501 serum specimens from unselected patients with invasive cervical cancer, 52.9% reacted positively in at least one of the four assays. In contrast, among 244 serum specimens from control subjects without cervical cancer, only 2 reactive serum specimens (0.8%) were found. For 19 of 19 antibody-positive patients, the HPV type indicated by seroreactivity was identical to the HPV DNA type found in the tumor, which also indicates a high degree of specificity for antibody detection with respect to HPV type. In a direct comparison of 72 serum specimens from patients with cervical cancer, 56% of the specimens reacted in at least one of the four protein ELISAs, whereas 40% reacted in at least one of seven peptide ELISAs covering the four antigens. These assays could be of value for the detection of invasive cervical cancer in settings in which cytology-based early tumor screening is not available, for the clinical management of patients diagnosed with cervical cancer, and for the immunological monitoring of E6 and E7 vaccination trials.
- Published
- 1998
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164. Priming of cytotoxic T lymphocytes by five heat-aggregated antigens in vivo: conditions, efficiency, and relation to antibody responses.
- Author
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Speidel K, Osen W, Faath S, Hilgert I, Obst R, Braspenning J, Momburg F, Hämmerling GJ, and Rammensee HG
- Subjects
- Animals, Antibody Formation, Antigens, Viral immunology, H-2 Antigens immunology, Hot Temperature, Immunity, Cellular, Mice, Mice, Inbred BALB C, Mice, Inbred C57BL, Mice, Transgenic, Oncogene Proteins, Viral immunology, Ovalbumin chemistry, Papillomaviridae chemistry, Papillomavirus E7 Proteins, Protein Denaturation, Vaccines immunology, beta-Galactosidase chemistry, Ovalbumin immunology, Papillomaviridae immunology, T-Lymphocytes, Cytotoxic immunology, beta-Galactosidase immunology
- Abstract
Mice were immunized i.p. with soluble or heat-denatured protein antigens [ovalbumin, beta-galactosidase, or recombinant E7 protein of human papilloma virus type 16 (HBV)]. Heat-denatured (100 degrees C) preparations of these proteins were able to induce cytotoxic T lymphocytes (CTL) that recognize cells expressing the respective genes, whereas native protein was either inefficient or required up to 30-fold higher doses. If the heat-treated proteins were separated into aggregated and soluble fractions by ultracentrifugation, only the aggregated fractions were able to induce specific CTL; this is probably because of the easier access to one of the major histocompatibility complex class I loading pathways for exogenous antigen. Addition of the adjuvant aluminium hydroxide (alum) to aggregated proteins abolished their ability to induce CTL; thus, a condition leading to a strong antibody response appeared to inhibit CTL induction. Interestingly, immunization with heat-denatured ovalbumin plus alum increased the IgM/IgG1 ratio compared to immunization with native ovalbumin and alum. Immunization of B6 mice transgenic for an HLA-A2/H-2K(b) hybrid gene with heat-denatured, recombinant HPV 16-E7 protein induced D(b)-restricted CTL specific for the peptide 49-57 of E7, indicating that this epitope is immunodominant over any A2-restricted E7 epitope in these mice. A whole influenza virus preparation heated to 100 degrees C or even autoclaved was still able to induce virus-specific CTL and BALB/c spleen cells heated to 100 degrees C could still cross-prime minor H-specific CTL in B6 mice, although with lower efficiency than fresh spleen cells. Thus, aggregated proteins can be considered as components for future vaccines.
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- 1997
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165. Chimeric papillomavirus-like particles.
- Author
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Müller M, Zhou J, Reed TD, Rittmüller C, Burger A, Gabelsberger J, Braspenning J, and Gissmann L
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- Amino Acid Sequence, Animals, Antibodies, Viral biosynthesis, Cell Line, Chlorocebus aethiops, Humans, Immunization, Mice, Mice, Inbred BALB C, Molecular Sequence Data, Oncogene Proteins, Viral genetics, Oncogene Proteins, Viral metabolism, Papillomaviridae immunology, Papillomaviridae metabolism, Papillomaviridae physiology, Papillomavirus E7 Proteins, Spodoptera cytology, Virion, Virus Assembly, Capsid Proteins, Papillomaviridae genetics, Recombination, Genetic
- Abstract
We have constructed chimeric papillomavirus-like particles (CVLPs) by replacing the 34-carboxy-terminal amino acids of the HPV 16 L1 protein with various parts of the HPV 16 E7 protein. Chimeric proteins were expressed by recombinant baculoviruses and analyzed by electron microscopy for their ability to assemble into virus capsids. We were able to produce CVLPs in high efficiencies with inserts of up to 60 amino acids. CVLPs are able to induce a neutralizing antibody response, assayed by inhibition of hemagglutination of mouse erythrocytes. CVLPs are interacting with the putative receptor for papillomaviruses as they were shown to hemagglutinate mouse red blood cells and bind to and penetrate cells in vitro. As CVLPs follow a similar intracellular pathway as observed earlier for BPV VLPs, we speculate that CVLPs can be used to deliver peptides into mammalian cells in vitro and in vivo, possibly reaching the pathway for MHC class I presentation.
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- 1997
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166. A general purification protocol for E7 proteins from "high- and low-risk" human papillomavirus types expressed in the yeast Schizosaccharomyces pombe.
- Author
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Braspenning J, Manetti R, Zumbach K, Meschede W, Gissmann L, and Tommasino M
- Subjects
- Animals, DNA biosynthesis, Female, Humans, Mice, Oncogene Proteins, Viral biosynthesis, Papillomaviridae pathogenicity, Papillomavirus E7 Proteins, Recombinant Fusion Proteins chemistry, Recombinant Fusion Proteins isolation & purification, Recombinant Fusion Proteins metabolism, Schizosaccharomyces virology, Condylomata Acuminata virology, Oncogene Proteins, Viral genetics, Oncogene Proteins, Viral isolation & purification, Papillomaviridae chemistry, Papillomavirus Infections virology, Schizosaccharomyces genetics, Tumor Virus Infections virology, Uterine Cervical Neoplasms virology
- Abstract
A purification protocol was developed to obtain human papillomavirus (HPV) type 16 E7 protein expressed in the yeast Schizosaccharomyces pombe. Only three chromatographic steps were necessary to purify the unfused HPV 16 E7 protein to homogeneity (95-99%) as shown by silver staining after polyacrylamide gel electrophoresis. Approximately 0.8 mg of highly purified E7 was obtained from 5 x 10(10) yeast cells. The purified HPV 16 E7 phosphoprotein (Ser 31/32) was refolded and assayed for functionality. Binding to the proteins Rb1 and p107 in vitro and induction of DNA synthesis after microinjection into serum-deprived NIH 3T3 cells suggest that the E7 protein retains some of its biological activities. Most importantly, the purification strategy is also applicable for different HPV 16 E7 mutants and for E7 proteins from other HPV types such as HPV 18 and 11.
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- 1997
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167. The human papillomavirus type 16 E7 protein is associated with the nucleolus in mammalian and yeast cells.
- Author
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Zatsepina O, Braspenning J, Robberson D, Hajibagheri MA, Blight KJ, Ely S, Hibma M, Spitkovsky D, Trendelenburg M, Crawford L, and Tommasino M
- Subjects
- Antibodies, Monoclonal, Cell Cycle physiology, Female, Humans, Immunohistochemistry, Microscopy, Electron, Microscopy, Fluorescence, Retinoblastoma Protein analysis, Schizosaccharomyces ultrastructure, Tumor Cells, Cultured, Uterine Cervical Neoplasms pathology, Uterine Cervical Neoplasms ultrastructure, Cell Nucleolus metabolism, Oncogene Proteins, Viral metabolism, Repressor Proteins, Schizosaccharomyces metabolism, Uterine Cervical Neoplasms metabolism
- Abstract
In this study we show, by immunofluorescence and electron microscopy immuno-gold labelling, that the major transforming protein of Human Papillomavirus type 16 E7 is associated with the nucleolus of cells derived from the HPV16-positive cervical carcinoma line CaSki. The E7 nucleolar staining appeared to be cell cycle dependent, being considerably reduced in the G2 phase. The total level of the protein in the cell, however, remained constant during all phases. We also show that the cellular protein Rb1, which is targeted by E7, is localised in the nucleus and nucleolus in CaSki cells. Thus, it is possible that the presence of E7 in the nucleolus correlates with a hypothetical function(s) of Rb1 in this particular intranuclear compartment. The nucleolar localisation of HPV16 E7 protein was also observed in the fission yeast Schizosaccharomyces pombe, suggesting that a targeting mechanism of HPV16 E7 protein into the nucleolus is common to both mammalian and yeast systems. Nucleolar localisation of HPV16 E7 protein may be independent from Rb1 since no Rb1 related proteins have been identified in fission yeast.
- Published
- 1997
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168. p53-independent growth regulation of cervical cancer cells by the papillomavirus E6 oncogene.
- Author
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Spitkovsky D, Aengeneyndt F, Braspenning J, and von Knebel Doeberitz M
- Subjects
- Amino Acid Sequence, Carcinoma genetics, Carcinoma pathology, Carcinoma virology, Cell Cycle genetics, Cell Division drug effects, Cell Division genetics, Clone Cells, Cyclin-Dependent Kinase Inhibitor p21, Cyclins genetics, Dexamethasone pharmacology, Female, G1 Phase genetics, Gene Expression Regulation, Neoplastic, Gene Expression Regulation, Viral, Genes, Reporter, Humans, Molecular Sequence Data, Mutation, Oncogene Proteins, Viral biosynthesis, Plasmids genetics, Trans-Activators, Transcription, Genetic, Tumor Cells, Cultured, Tumor Suppressor Protein p53 metabolism, Uterine Cervical Neoplasms pathology, Uterine Cervical Neoplasms virology, DNA-Binding Proteins, Oncogene Proteins, Viral genetics, Repressor Proteins, Tumor Suppressor Protein p53 genetics, Uterine Cervical Neoplasms genetics
- Abstract
Growth of cervical carcinoma cells depends on continuous expression of high risk type human papillomavirus oncogenes E6 and E7. E6 destabilizes p53, a tumor-suppressive transcription factor, which activates expression of the inhibitor of cell cycle progression p21 and other genes. E6-mediated p53 degradation can therefore result in cell cycle deregulation. It has, however, not yet been determined whether p53 inactivation is sufficient to provoke cell cycle progression in established cervical carcinoma cells. Moreover, it has not yet been clarified whether E6 confers additional p53-independent growth stimuli in cancer cells. To address these questions, we analysed p53 functions in SW 756 cervical cancer cells in which the expression of endogenous HPV 18 E6-E7 genes can be downregulated by dexamethasone. This results in significantly increased p53 levels and subsequent cell cycle arrest in the Gz phase. Surprisingly, p53 activities were suppressed rather than enhanced in these cervical cancer cells. However, if high risk papillomavirus type 16 E6 genes, including a mutant which does not degrade p53, were expressed in dexamethasone-treated SW 756 cells with suppressed endogenous HPV type 18 E6-E7 expression, the cells reentered the cell cycle even in the absence of a cooperating viral E7 gene. In contrast, the non oncogenic papillomavirus type 6 E6 gene did not release the cells from growth arrest under these conditions. These data indicate that suppression of p53 functions is not sufficient to provoke cell cycle progression in E6-E7-depleted cervical cancer cells and point to a p53-independent mitotic activity to oncogenic papillomavirus type E6 genes in cervical carcinoma cells.
- Published
- 1996
169. Quality of life after acoustic neuroma surgery.
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van Leeuwen JP, Braspenning JC, Meijer H, and Cremers CW
- Subjects
- Adult, Aged, Cranial Nerve Neoplasms pathology, Ear, Inner surgery, Employment, Facial Nerve physiopathology, Female, Health Services statistics & numerical data, Health Status, Humans, Male, Middle Aged, Neuroma, Acoustic pathology, Occipital Lobe surgery, Patient Satisfaction, Retrospective Studies, Social Class, Treatment Outcome, Vestibulocochlear Nerve pathology, Cranial Nerve Neoplasms surgery, Neuroma, Acoustic surgery, Quality of Life, Vestibulocochlear Nerve surgery
- Abstract
A questionnaire was sent to 134 patients who had undergone surgery for a unilateral acoustic neuroma between 1980 and 1993, to obtain data on the consequences on their quality of life, physical condition, social life, employment, and use of medical facilities. Distinctions were made between the translabyrinthine-transotic approach, the suboccipital approach, the tumor size, and the number of operations per patient. We found that the patients' reported state of health after surgery was poorer than that in a group of comparable nonoperated patients. Recuperation after an operation took many months and did not always result in full recovery. Surgery had various effects on preoperative symptoms such as hearing loss, tinnitus, vertigo, and facial nerve dysfunction: improvement, no change, or deterioration. Surgery had severe consequences on social life and occupation, but far less effect on income. Almost one third of the patients required postoperative home help, and a proportion were declared unfit to work. The surgical approach, tumor size, and reoperations had a definite influence on the study parameters. After suboccipital surgery, there were more reports of pain, more declarations of incapacity to work, poorer facial nerve function, and more frequent visits to the general practitioner. The translabyrinthine-transotic approach was associated with more severe pain and more complaints of postoperative vertigo. A greater proportion of the patients with larger tumors were declared unfit to work. The general state of health after suboccipital reoperations was better than after the initial operation; there was no reasonable explanation for this. Facial nerve function deteriorated after reoperation(s).
- Published
- 1996
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