Endovascular Cerebral Venous Sinus Imaging with Optical Coherence Tomography.

BACKGROUND AND PURPOSE: Imaging of the cerebral venous sinuses has evolved Substantially during the past 2 decades, and most recently intravascular sinus imaging with sonography has shed light on the pathophysiology of sinus thrombosis and intracranial hypertension. Optical coherence tomography is the highest resolution intravascular imaging technique available but has not been previously used in cerebral sinus imaging. The purpose of this study was to develop a preclinical animal model of endovascular optical coherence tomography cerebral venous sinus imaging and compare optical coherence tomography findings with histology. MATERIALS AND METHODS: Four consecutive Yorkshire swine were selected. The superior sagittal sinus was first catheterized with a microwire, and the optical coherence tomography catheter was delivered via a monorail technique into the sinus. Luminal blood was cleared with a single arterial injection. After structural and Doppler optical coherence tomography imaging, a craniotomy was performed and the sinus and adjacent dura/veins were resected. Bland-Altman analysis was performed to compare optical coherence tomography and histology. RESULTS: Technically successful optical coherence tomography images were obtained in 3 of 4 swine. The luminal environment and visualization of dural arteries and draining cortical veins were characterized. The average maximum diameters of the sinus, dural arteries, and cortical veins were 3.14 mm, 135 µm, and 260 µm, respectively. Bland-Altman analysis demonstrated good agreement between histology and optical coherence tomography images. CONCLUSIONS: Endovascular optical coherence tomography imaging was feasible in this preclinical animal study. Adoption of this imaging technique in the human cerebral venous sinus could aid in the diagnosis, treatment, and understanding of the pathophysiology of various diseases of the sinus. Human safety and feasibility studies are needed.

18F-FDG PET/CT, cytoreductive surgery and intraperitoneal chemohyperthermia for the therapeutic management in peritoneal carcinomatosis: a pilot study / 18F-FDG PET/TC, cirugía citorreductora y quimiohipertermia intraperitoneal para el tratamiento terapéutico de la carcinomatosis peritoneal: estudio piloto

Aim. Peritoneal carcinomatosis is a common evolution of neoplasms and the terminal stage of disease. A new therapeutic technique, based on the total surgical removal of peritoneal lesions (peritonectomy procedure - PP) combined with the intraperitoneal chemohyperthermia (IPCH), has been developed. Proper patient selection is mandatory for optimizing the results of treatment. The aim of this study was to investigate the role of [(18)F]fluoro-2-deoxy-d-glucose Positron Emission Tomography/Computed Tomography (18F-FDG PET/CT) in patients with peritoneal carcinosis selected to undergo PP and IPCH. Furthermore, we aimed to identify characteristic patterns of abdominal18F-FDG uptake and to correlate these patterns with available anatomic findings after surgery. Methods. Patients with either histologically confirmed peritoneal carcinosis or suspected upon clinical follow-up and/or imaging findings were prospectively submitted to pre-surgery 18F-FDG PET/CT scan. Only those patients without evidence of extra-peritoneal metastases at PET/CT scan were treated with PP and IPCH. Results. 11 patients with peritoneal carcinomatosis (5 colorectal, 4 ovarian, 1 pancreatic) and 1 unknown primitive cancer, were eligible for the study. In all cases PET/CT scan showed multiple peritoneal implants. In 6 out of 11 cases (54%) metastases were evidenced by 18F-FDG PET/CT: 2 cases with liver metastases; 1 case with bone metastases; 3 patients with lymph-node lesions. Two distinct imaging patterns, with focal or diffuse increased 18F-FDG uptake, were recognized. Conclusions. PP + IPCH of patients selected by 18F-FDG PET/CT seems to be safe and feasible. PET/CT scan appears as a reliable tool for the detection, characterization of peritoneal implants with potential impact in the therapeutic management of these patients (AU)

Objetivo. La carcinomatosis peritoneal es una evolución común de las neoplasias y constituye el estadio terminal de la enfermedad. Se ha desarrollado una nueva técnica, basada en la extirpación quirúrgica de las lesiones peritoneales (procedimiento de peritonectomía - PP), combinada con quimiohipertermia intraperitoneal (IPCH). La adecuada selección de los pacientes es primordial, a fin de optimizar los resultados del tratamiento. El objetivo de este estudio fue investigar el papel de la tomografía de emisión de positrones con [(18)F]fluoro-2-deoxy-d-glucosa/tomografía computarizada (18F-FDG PET/TC) en pacientes con carcinomatosis peritoneal, seleccionados para someterse a PP e IPCH. Además, tratamos de identificar los patrones característicos de la captación abdominal de 18F-FDG y correlacionar dichos patrones con los hallazgos anatómicos disponibles tras la cirugía. Métodos. Se realizaron exámenes 18F-FDG PET/TC de manera prospectiva, y previamente a la cirugía, a los pacientes con carcinomatosis peritoneal histológicamente confirmada, o sospechada mediante seguimiento clínico y/o hallazgos de imagen. Solo puede tratarse con PP y IPCH a aquellos pacientes que no reflejen evidencia de metástasis extraperitoneales en los exámenes PET/TC. Resultados. Se seleccionó para el estudio a 11 pacientes con carcinomatosis peritoneal (5 colorrectales, 4 ováricas, una pancreática) y un cáncer primitivo desconocido. En todos los casos, el examen PET/TC reflejó múltiples implantes peritoneales. En 6 de los 11 casos (54%) las metástasis fueron evidenciadas mediante 18F-FDG PET/TC: 2 casos con metástasis hepáticas, un caso con metástasis óseas, y 3 pacientes con lesiones ganglionares. Se reconocieron 2 patrones de imagen distintos, con aumento de captación focal o difusa de 18F-FDG. Conclusiones. La combinación PP + IPCH de los pacientes seleccionados mediante 18F-FDG PET/TC parece ser una técnica segura y factible. La PET/TC se revela como una herramienta fiable para la detección y caracterización de los implantes peritoneales, con un impacto potencial sobre el tratamiento terapéutico de dichos pacientes (AU)

(18)F-FDG PET/CT, cytoreductive surgery and intraperitoneal chemohyperthermia for the therapeutic management in peritoneal carcinomatosis: A pilot study.

AIM: Peritoneal carcinomatosis is a common evolution of neoplasms and the terminal stage of disease. A new therapeutic technique, based on the total surgical removal of peritoneal lesions (peritonectomy procedure - PP) combined with the intraperitoneal chemohyperthermia (IPCH), has been developed. Proper patient selection is mandatory for optimizing the results of treatment. The aim of this study was to investigate the role of [(18)F]fluoro-2-deoxy-d-glucose Positron Emission Tomography/Computed Tomography ((18)F-FDG PET/CT) in patients with peritoneal carcinosis selected to undergo PP and IPCH. Furthermore, we aimed to identify characteristic patterns of abdominal(18)F-FDG uptake and to correlate these patterns with available anatomic findings after surgery. METHODS: Patients with either histologically confirmed peritoneal carcinosis or suspected upon clinical follow-up and/or imaging findings were prospectively submitted to pre-surgery (18)F-FDG PET/CT scan. Only those patients without evidence of extra-peritoneal metastases at PET/CT scan were treated with PP and IPCH. RESULTS: 11 patients with peritoneal carcinomatosis (5 colorectal, 4 ovarian, 1 pancreatic) and 1 unknown primitive cancer, were eligible for the study. In all cases PET/CT scan showed multiple peritoneal implants. In 6 out of 11 cases (54%) metastases were evidenced by (18)F-FDG PET/CT: 2 cases with liver metastases; 1 case with bone metastases; 3 patients with lymph-node lesions. Two distinct imaging patterns, with focal or diffuse increased (18)F-FDG uptake, were recognized. CONCLUSIONS: PP+IPCH of patients selected by (18)F-FDG PET/CT seems to be safe and feasible. PET/CT scan appears as a reliable tool for the detection, characterization of peritoneal implants with potential impact in the therapeutic management of these patients.

Comparison of CT and chemical-shift MRI for differentiating thymoma from non-thymomatous conditions in myasthenia gravis: value of qualitative and quantitative assessment.

AIM: To evaluate the usefulness of computed tomography (CT) and chemical-shift magnetic resonance imaging (MRI) in patients with myasthenia gravis (MG) for differentiating thymoma (THY) from thymic lymphoid hyperplasia (TLH) and normal thymus (NT), and to determine which technique is more accurate. MATERIALS AND METHODS: Eighty-three patients with generalised MG who underwent surgery were divided into the TLH/NT group (A; 65 patients) and THY group (B; 24 patients). Differences in qualitative characteristics and quantitative data (CT: radiodensity in Hounsfield units; MRI: signal intensity index [SII]) between groups were tested using Fisher's exact test and Student's t-test. Logistic regression models were estimated for both qualitative and quantitative analyses. At quantitative analysis, discrimination abilities were determined according to the area under the receiver operating characteristic (ROC) curve (AUROC) with computation of optimal cut-off points. The diagnostic accuracies of CT and MRI were compared using McNemar's test. RESULTS: At qualitative assessment, MRI had higher accuracy than CT (96.4%, 80/83 and 86.7%, 72/83, respectively). At quantitative analysis, both the radiodensity and SII were significantly different between groups (p<0.0001). For CT, at quantitative assessment, the AUROC of the radiodensity in discriminating between groups was 0.904 (optimal cut-off point, 20 HU) with an accuracy of 77.1% (64/83). For MRI, the AUROC of the SII was 0.989 (optimal cut-off point, 7.766%) with an accuracy of 96.4% (80/83), which was significantly higher than CT (p<0.0001). By using optimal cut-off points for cases with an erroneous diagnosis at qualitative assessment, accuracy improved both for CT (89.2%, 74/83) and MRI (97.6%, 81/83). CONCLUSION: Quantitative analysis is useful in evaluating patients with MG and improves the diagnostic accuracy of CT and MRI based on qualitative assessment. Chemical-shift MRI is more reliable than CT in differentiating THYs from non-thymomatous conditions.

Clinical management of critically ill patients with Cushing's disease due to ACTH-secreting pituitary macroadenomas: effectiveness of presurgical treatment with pasireotide.

The management of critically ill Cushing's disease (CD) patients is extremely challenging. Pasireotide is indicated for the treatment of CD patients when pituitary surgery is unfeasible or has not been curative, but no data are available about the use of this drug as pre-operative treatment in critically ill patients. We report the effects of presurgical pasireotide therapy in CD patients in whom hypercortisolism caused life-threatening hypokalemia, alkalosis, and cardio-respiratory complications precluding surgical approach. Clinical, biochemical, and radiological data of two critically ill patients with ACTH-secreting pituitary macroadenoma, before and during first-line presurgical pasireotide treatment (600 µg s.c. bid). During the first 21 days of treatment, pasireotide therapy induced a rapid, partial decrease of plasma ACTH, serum cortisol, and urinary free cortisol levels, with the consequent normalization of serum potassium concentration and arterial blood gases parameters, in both the patients. They did not experience unmanageable side effects and underwent endoscopic transsphenoidal surgery after 4 weeks of effective treatment. Pre-operative MRI evaluation did not show pituitary tumor shrinkage. Surgical cure of CD was obtained in the first patient, while debulking allowed the pharmacological control of hypercortisolism in the second case. We suggest that pasireotide can induce a rapid improvement of clinical and metabolic conditions in critically ill CD patients in whom surgical approach is considered hazardous and need to be delayed.

Imaging of thymus in myasthenia gravis: from thymic hyperplasia to thymic tumor.

Myasthenia gravis (MG) is an autoimmune disorder often associated with thymic abnormalities. At onset, thymic lymphoid hyperplasia (TLH) and thymoma can be found in up to 65% and 15% of patients, respectively. Diagnostic imaging is crucial in this setting in order to detect the presence and type of the thymic abnormality and in the preoperative planning, when indicated. Chest radiography has a minor role due to its low accuracy. Computed tomography is the imaging modality of choice, although the differentiation between a small thymoma and TLH that appears as a focal soft-tissue mass may be not possible. Magnetic resonance imaging (MRI) is not usually employed, but it is useful in equivocal cases, especially in differentiating focal TLH from thymoma by using chemical-shift sequences for defining the proper management. In addition, diffusion-weighted (DW)-MRI can differentiate lipid-poor normal/hyperplastic thymus from thymoma and could be useful in differentiating non-advanced from advanced thymomas. Positron emission tomography (PET)-CT is not helpful in distinguishing early from advanced thymoma but can be used to differentiate thymic carcinoma from thymoma. Hereby, we discuss the imaging features of thymic abnormalities in MG, even focusing on novel aspects of chemical-shift and DW-MRI.

Late presentation of ureteral injury following laparoscopic colorectal surgery.

Iatrogenic ureteral injury is an uncommon but dangerous complication of abdominal and pelvic surgery. When recognized and promptly treated, most ureteral lesions heal without sequelae. Instead, undetected injuries may last for a prolonged period of time since symptoms and signs are usually subtle and nonspecific, even if evolution may be life threatening. In doubtful cases the diagnostic role of modern multiphase helical computed tomography is crucial. We describe the late presentation in the Emergency Department of a case of double left ureteral injury after abdominal surgery, and illustrate the appearance of the lesions at computerized tomography.

Computed tomography and thymoma: distinctive findings in invasive and noninvasive thymoma and predictive features of recurrence.

PURPOSE: Our goal was to assess the computed tomography (CT) imaging findings of thymoma and to correlate these features with Masaoka staging system and prognosis. MATERIALS AND METHODS: CT findings of thymoma were analysed in 58 patients who had undergone surgery between January 2002 and September 2007. All cases were classified according to the Masaoka staging system. The presence of various CT findings was correlated with tumour invasiveness and recurrence. In statistical analysis, a p value <0.05 was interpreted as significant. RESULTS: The study found 26 noninvasive thymomas and 32 invasive thymomas. Invasive thymomas were more likely to be greater in size (p<0.01), with lobulated or irregular contours (p<0.02), a necrotic or cystic component (p<0.04), foci of calcification (p<0.05) and heterogeneous contrast enhancement (p<0.01) than were noninvasive thymomas. Disease progression developed in nine of 58 patients. Tumour recurrence and metastasis correlated with greater size (p<0.04), lobulated or irregular contours (p<0.01), complete mediastinal fat obliteration (p<0.01), great vessel invasion (p<0.01) and pleural implants (p<0.02). CONCLUSIONS: CT is useful in differentiating invasive from noninvasive thymomas and plays an important role in evaluating and treating these patients for multimodal therapy with neoadjuvant approaches. Moreover, CT findings may serve as predictors of postoperative recurrence or metastasis.

Clinical application of lung ultrasound in patients with acute dyspnea: differential diagnosis between cardiogenic and pulmonary causes.

This review discusses the usefulness of bedside lung ultrasound in the diagnostic distinction between the various causes of acute dyspnoea in the emergency department, with special attention to the differential diagnosis of pulmonary oedema and exacerbation of chronic obstructive pulmonary disease (COPD). This is made possible by using mid- to low-end scanners and simple acquisition techniques accessible to both radiologists and clinicians. Major advantages include ready availability at the bedside, the absence of ionising radiation, high reproducibility and cost efficiency. The technique is based on the recognition and analysis of sonographic artefacts rather than direct visualisation of the pulmonary structures. These artefacts are caused by the interaction of water-rich structures and air, called comet tails or B-lines. When such artefacts are widely detected on anterolateral transthoracic lung scans, diffuse alveolar-interstitial syndrome can be diagnosed, which is often a sign of acute pulmonary oedema. This condition rules out exacerbation of COPD as the main cause of acute dyspnoea.

CT-guided percutaneous transthoracic biopsy in the diagnosis of mediastinal masses: evaluation of 73 procedures.

PURPOSE: This study was performed to evaluate the factors affecting the diagnostic accuracy and rate of complications of CT-guided percutaneous transthoracic needle biopsy of mediastinal masses. MATERIALS AND METHODS: We reviewed 73 consecutive mediastinal biopsies in 70 patients. Final diagnoses were based on a retrospective analysis of surgical outcomes, results of repeat biopsies or findings of imaging and clinical follow-up lasting at least 4 months. Benign and malignant biopsy findings were compared with the final outcomes to determine the diagnostic accuracy of the method. Finally, we analysed the complications. RESULTS: CT-guided percutaneous transthoracic needle biopsy provided adequate samples in 61/73 cases, with a total sample rate of 83.6%. Of these 61 biopsies, 51 yielded a correct diagnosis with specific histological typing, mainly in the case of thymoma and metastasis. Lymphomas were less reliably diagnosed. The overall sensitivity, specificity, positive predictive value, negative predictive value and diagnostic accuracy values were 83.6%, 100%, 100%, 35.3% and 83.6%, respectively. Pneumothorax was the most common complication (5.5%). CONCLUSIONS: CT-guided percutaneous transthoracic needle biopsy is an easy, reliable and safe procedure that obviates the need for exploratory surgery in medically treatable or unresectable cases. It should be the first invasive procedure in the diagnostic workup of mediastinal masses.

Accuracy of CT-guided transthoracic needle biopsy of lung lesions: factors affecting diagnostic yield.

PURPOSE: This study was performed to analyse the variables affecting the diagnostic accuracy of computed tomography (CT)-guided transthoracic needle biopsy of pulmonary lesions. MATERIALS AND METHODS: A retrospective study of 612 consecutive procedures with confirmed final diagnoses was undertaken. Benign and malignant needle biopsy results were compared with final outcomes to determine diagnostic accuracy. A statistical analysis of factors related to patient characteristics, lung lesions and biopsy technique was performed to determine possible influences on diagnostic yield. A p value less than 0.05 was interpreted as statistically significant. RESULTS: There were 508 (83%) malignant and 104 (17%) benign lesions. Sensitivity, specificity, positive predictive value, negative predictive value and accuracy for a diagnosis of malignancy were 90.2%, 99.0%, 99.8%, 67.3% and 91.7%, respectively. Overall diagnostic accuracy was 83.3%. Variables affecting diagnostic accuracy were the final diagnosis (benign 67%, malignant 92%; p<0.001) and lesion size (lesions<1.5 cm 68%, lesions 1.5-5.0 cm 87%, lesions>5 cm 78%; p<0.05). CONCLUSIONS: In CT-guided transthoracic needle biopsy, the final diagnosis and lesion size affect diagnostic accuracy: benign lung lesions and lesions smaller than 1.5 cm or larger than 5.0 cm in diameter provide lower diagnostic yield.

Contrast-enhanced ultrasonography in Crohn's disease.

PURPOSE: The purpose of this study was to demonstrate the diagnostic efficacy of contrast-enhanced power Doppler (CEPD) after ultrasound (US) with colour power Doppler (CPD) in defining disease activity in patients with ileal Crohn's disease in the acute phase and during treatment follow-up. MATERIALS AND METHODS: The study included 15 patients with ileal Crohn's disease, 12 of whom were examined in the acute phase of their illness and then 30-60 days after treatment. Three patients were studied during clinical quiescence. All patients were examined prior to treatment with US-CPD study and then with contrast-enhanced power Doppler (CEPD) examination (CEPD) with the use of US contrast agent (SonoVue, Bracco), together with clinical assessment and laboratory tests. Disease activity was defined by the Crohn's Disease Activity Index (CDAI) and some of the most sensitive biohumoural markers. After initial US, CPD and CEPD were performed to assess enhancement of the thickened bowel wall with the use of a reference box and a semiquantitative scoring system. RESULTS: The CEPD study is more reliable then US-CPD in the diagnosis of Crohn's disease and statistically agrees significantly with clinical and laboratory indices for disease activity. CONCLUSIONS: This study demonstrates the importance of US-CPD in the follow-up of patients with Crohn's disease and suggests systematic use of the US contrast agent, which can improve diagnostic performance of abdominal US study. It also provides more information about patients both in the acute phase and during follow-up, thus improving treatment planning and better monitoring of treatment efficacy.

The anterior mediastinum: anatomy and imaging procedures.

The mediastinum is divided into compartments (anterior, middle, posterior) on the basis of lateral chest radiographs. Several anatomical and radiological classifications of the mediastinum are reported in the literature. Most mediastinal abnormalities are initially suspected following chest radiography; the need for further investigation and the most appropriate imaging modality are largely dictated by the tentative diagnosis made on this examination. Although routine chest radiography initiates the evaluation of mediastinal disorders, it is rarely diagnostic: notable exceptions are teeth or bones within a mass, which are diagnostic of a teratoma; air/fluid levels suggest an oesophageal origin, hernia, cyst, or abscess. Chest radiography is followed by spiral computed tomography (sCT). However, even sCT with contrast material is occasionally diagnostic (a confident diagnosis can be made of some lesions such as mature teratoma and mediastinal goiter) but is usually sufficient for preoperative evaluation before mediastinotomy or mediastinoscopy: it is instrumental in planning further diagnostic workup. In certain cases, magnetic resonance imaging (MRI) may be complementary to sCT, but its use is not considered routine. Besides, although the anterior mediastinum is suitable for sonographic examination, the diagnostic value of ultrasonography has not been fully exploited. Thyroid scanning with radioactive iodine is useful in identifying and evaluating masses of suspected thyroid origin. The role of fluorodeoxyglucose positron emission tomography (FDG-PET) in mediastinal diseases continues to be evaluated: it has potential for differentiating between benign and malignant disease and is expected to play a more extensive role in the imaging of mediastinal neoplasms in the future. In this paper, the radiological features of masses located in the anterior mediastinum are discussed, with particular reference to radiographic and CT patterns useful to the clinician's everyday practice.

The anterior mediastinum: diseases.

Mediastinal tumours are frequently asymptomatic and first noted on routine chest radiograph. In most cases, evaluation should proceed to spiral computed tomography (sCT) of the chest with iodinated contrast material. The specific location and appearance of tumours on sCT is instrumental in planning further diagnostic and treatment strategies. Primary tumours in the anterior mediastinum account for half of all mediastinal masses. They comprise various benign and malignant neoplasms, but a wide variety of nonneoplastic lesions (developmental, inflammatory) can present as a localised mass in this compartment. The most common primary anterior mediastinal tumours are thymoma, teratoma and lymphoma; all other lesions are rare. Nonneoplastic conditions include thymic cysts, lymphangioma and intrathoracic goitre. Understanding the pathology, clinical presentation, imaging and diagnosis of the major tumour types is instrumental in the safe and efficient work-up of a mediastinal mass. Patients with primary mediastinal masses and cysts will usually undergo surgical resection; radiological and clinical features should prompt limited biopsy specimens followed by oncologic consultation, and chemotherapy or radiotherapy when appropriate. The objective of this review was to examine the role of diagnostic imaging in the management of masses of the anterior mediastinum.

Species barriers in prion diseases--brief review.

Transmissible spongiform encephalopathies (TSEs or prion diseases) are neurological disorders associated with the aggregation of a pathologic isoform of a host-encoded protein, termed prion protein (PrP). The pathologic isoform of PrP, termed PrP(Sc), is a major constituent of the infectious agent. TSE diseases are characterized by neurodegenerative failure and inevitable morbidity. Bovine spongiform encephalopathy (BSE) has been transmitted from cattle to humans to cause a new variant of Creutzfeldt-Jakob syndrome. The potential for chronic wasting disease to similarly cross the species barrier from cervids to humans is considered unlikely but possible. Thus, understanding how TSE agents overcome resistance to transmission between species is crucial if we are to prevent future epidemics. The species barrier usually can be abrogated to varying degrees in laboratory animals. Studies done with transgenic animals, tissue culture, and cell-free assays established PrP as being necessary for TSE pathogenesis and illustrated that certain amino acid residues are more influential than others for conferring resistance to TSE agent transmission. The essence of what constitutes a TSE agent's species compatibility is thought to be orchestrated by a complex interplay of contributions from its primary amino acid sequence, its glycoform patterns, and its three-dimensional structure.

Glycosylation influences cross-species formation of protease-resistant prion protein.

A key event in the transmissible spongiform encephalopathies (TSEs) is the formation of aggregated and protease-resistant prion protein, PrP-res, from a normally soluble, protease-sensitive and glycosylated precursor, PrP-sen. While amino acid sequence similarity between PrP-sen and PrP-res influences both PrP-res formation and cross-species transmission of infectivity, the influence of co- or post-translational modifications to PrP-sen is unknown. Here we report that, if PrP-sen and PrP-res are derived from different species, PrP-sen glycosylation can significantly affect PrP-res formation. Glycosylation affected PrP-res formation by influencing the amount of PrP-sen bound to PrP-res, while the amino acid sequence of PrP-sen influenced the amount of PrP-res generated in the post-binding conversion step. Our results show that in addition to amino acid sequence, co- or post-translational modifications to PrP-sen influence PrP-res formation in vitro. In vivo, these modifications might contribute to the resistance to infection associated with transmission of TSE infectivity across species barriers.

Deletion of beta-strand and alpha-helix secondary structure in normal prion protein inhibits formation of its protease-resistant isoform.

A fundamental event in the pathogenesis of transmissible spongiform encephalopathies (TSE) is the conversion of a normal, proteinase K-sensitive, host-encoded protein, PrP-sen, into its protease-resistant isoform, PrP-res. During the formation of PrP-res, PrP-sen undergoes conformational changes that involve an increase of beta-sheet secondary structure. While previous studies in which PrP-sen deletion mutants were expressed in transgenic mice or scrapie-infected cell cultures have identified regions in PrP-sen that are important in the formation of PrP-res, the exact role of PrP-sen secondary structures in the conformational transition of PrP-sen to PrP-res has not yet been defined. We constructed PrP-sen mutants with deletions of the first beta-strand, the second beta-strand, or the first alpha-helix and tested whether these mutants could be converted to PrP-res in both scrapie-infected neuroblastoma cells (Sc(+)-MNB cells) and a cell-free conversion assay. Removal of the second beta-strand or the first alpha-helix significantly altered both processing and the cellular localization of PrP-sen, while deletion of the first beta-strand had no effect on these events. However, all of the mutants significantly inhibited the formation of PrP-res in Sc(+)-MNB cells and had a greatly reduced ability to form protease-resistant PrP in a cell-free assay system. Thus, our results demonstrate that deletion of the beta-strands and the first alpha-helix of PrP-sen can fundamentally affect PrP-res formation and/or PrP-sen processing.

N-terminal truncation of prion protein affects both formation and conformation of abnormal protease-resistant prion protein generated in vitro.

Transmissible spongiform encephalopathy diseases are characterized by conversion of the normal protease-sensitive host prion protein, PrP-sen, to an abnormal protease-resistant form, PrP-res. In the current study, deletions were introduced into the flexible tail of PrP-sen (23) to determine if this region was required for formation of PrP-res in a cell-free assay. PrP-res formation was significantly reduced by deletion of residues 34-94 relative to full-length hamster PrP. Deletion of another nineteen amino acids to residue 113 further reduced the amount of PrP-res formed. Furthermore, the presence of additional proteinase K cleavage sites indicated that deletion to residue 113 generated a protease-resistant product with an altered conformation. Conversion of PrP deletion mutants was also affected by post-translational modifications to PrP-sen. Conversion of unglycosylated PrP-sen appeared to alter both the amount and the conformation of protease-resistant PrP-res produced from N-terminally truncated PrP-sen. The N-terminal region also affected the ability of hamster PrP to block mouse PrP-res formation in scrapie-infected mouse neuroblastoma cells. Thus, regions within the flexible N-terminal tail of PrP influenced interactions required for both generating and disrupting PrP-res formation.