Synovial cysts of the lumbar spine.

BACKGROUND AND PURPOSE: Synovial cysts of the spine occur most frequently in the lumbosacral region. Methods of treatment vary, but in cases of chronic pain or neurological deficits surgical intervention is undertaken. The aim of this paper is to present indications, surgical technique and efficacy of surgical treatment in patients with synovial cyst of the spinal canal. MATERIAL AND METHODS: The retrospective analysis included 11 patients, aged from 47 to 72 years, treated at the Department of Neurosurgery and Neurotraumatology, Poznan University of Medical Sciences, between 2004 and 2009. The length of medical history ranged from 2 months to 6 years. Conservative treatment applied before surgery was not effective. Neurological examination revealed unilateral or bilateral sciatica, superficial sensory disturbance or lower limb paresis. RESULTS: Synovial cysts were located mainly at the L4-L5 level (9 cases). Magnetic resonance imaging (MRI) of the spine was performed in all patients and showed the cystic lesion attached to the intervertebral joint. Surgical treatment consisted of a unilateral fenestration using microsurgical techniques in most cases. Back pain relief was observed in 9 cases. In 10 patients, symptoms of sciatica disappeared. Neurological deficits disappeared in 5 patients. CONCLUSIONS: Surgical treatment of spinal synovial cysts is safe, effective and ensures a long-lasting effect. Surgical treatment is indicated in patients in whom the clinical symptoms correlate with the presence of synovial cyst in imaging studies and do not resolve after conservative treatment.

Surgical treatment of symptomatic vertebral haemangiomas.

BACKGROUND AND PURPOSE: Vertebral haemangiomas are relatively common, benign vascular lesions; symptomatic ones that cause spinal cord compression are rare, however. Only 0.9-1.2% of all vertebral haemangiomas are symptomatic. The aim of the paper is to present indications, operative techniques and stabilization methods in patients with symptomatic vertebral haemangiomas. MATERIAL AND METHODS: Clinical analysis included 7 patients treated between 1995 and 2007. There were 4 females and 3 males, aged 24 to 63 yrs (average age 44 yrs). Symptomatic vertebral haemangiomas were diagnosed on the basis of neuroradiological studies. Surgery was applied in all cases. Implantation of internal stabilization followed vertebral haemangioma resection. RESULTS: Localization of vertebral haemangiomas included 1 case in the cervical, 5 cases in the thoracic and 1 case in the lumbar segment of the vertebral column. Symptoms of medulla compression were observed in 7 patients. Neurological symptoms were caused usually by hypertrophy or ballooning of the posterior cortex of the vertebral body into the vertebral canal. The anterior surgical approach was carried out in 2 cases, posterolateral in 3 cases and posterior in 2 cases. Spinal stability was secured by various implant systems and autogenic bone grafts. Bone defects in the vertebral body were filled with acrylic cement in 4 patients. In histological examinations, cavernous types were found in all patients. Neurological condition improved after the treatment in 5 patients. CONCLUSIONS: No standard therapy exists for symptomatic thoracic vertebral haemangiomas. However, immediate surgical intervention is necessary in cases with acute compressive myelopathy before the symptoms become irreversible.

Pituitary abscess: a case report.

OBJECTIVES: Pituitary abscess is rare disease and the correct diagnosis is difficult because there are non-specific symptoms and it is often radiologically indistinguishable from other pituitary lesions. CASE PRESENTATION: We present one case of pituitary abscess that constitute 0.15% of all pituitary adenomas operated in our department in the 20 years. A 49-year-old woman presented with a history of 10 months bifrontal headache. The MRI showed cystic intra and suprasellar mass with ring enhancement after contrast injection. During transsphenoidal surgery, copious yellowish pus was found. Antibiotic therapy was performed. Histological study of the cyst wall confirmed the diagnosis of pituitary abscess. CONCLUSION: Pituitary abscess should be considered in the differential diagnosis of all other cyst mass in patients with diabetes insipidus.

Application of internal stabilisation in the surgical treatment of spinal metastases.

BACKGROUND AND PURPOSE: Neoplastic disease damages the spine more often than trauma. Metastatic tumour causes vertebral column instability and neurological deficit. Surgical intervention indications depend on the patient's general and neurological status. The aim of neuro-orthopaedic treatment is to achieve pain relief and neurological improvement. Spine stability is secured by various implant systems. The aim of the paper is to present indications, operative techniques and stabilisation methods in patients with metastatic spine disease. MATERIAL AND METHODS: There were 73 patients included in this study: 50 males and 23 females, aged 17-74 years. The dominant tumour location was the thoracic spine (41 cases), followed by the lumbar spine (19). Symptoms of spinal cord lesion were observed in 71 patients. Qualification for surgery and approach planning were based on different scales (ASA, DeWald, Frankel, Karnofsky, Denis, Tomita). Internal stabilisation implantation followed tumour resection. RESULTS: Anterior approach was used in 15 cases, posterolateral in 39, posterior in 13, and combined in 6 cases. Histological findings generally allowed the primary tumour location to be disclosed, which in most cases involved the kidney, prostate, lung and the haematopoietic system. Neurological improvement was observed in 82% of cases. Perioperative death occurred in 3% of all patients. CONCLUSIONS: Employed operative techniques are adequate for tumour removal, neural and vascular structures decompression and for implant placement.

Evaluation of epidural scar formation in lumbar spine after TachoComb application - an experimental study.

BACKGROUND AND PURPOSE: Scar formation after spine surgery in the lumbosacral region may be the cause of failed back surgery syndrome. Therefore efforts are made to find materials preventing excessive scar formation at the site of surgery. The aim of the study was to evaluate the usefulness of TachoComb application in prevention of epidural scar formation in a rat experimental model. This paper additionally presents a review of literature concerning other methods of local suppression of scar formation after posterior approaches to the lumbar spine. MATERIAL AND METHODS: The experimental study was carried out on 14 male Wistar rats. Rats were divided into 2 groups. Laminectomy was performed in the first group (control group: n=5). In the second group of animals (n=9) laminectomy was followed by TachoComb application on the exposed dura. Neurological condition of the studied animals was evaluated based on clinical observation, neurological tests and recording of somatosensory evoked potentials. Post mortem histological examination was the main method of assessment of the experimental material. RESULTS: Presence of scar in the vertebral canal, its extent and severity differed between experimental groups. Electrophysiological results were also different between studied groups. CONCLUSIONS: TachoComb prevents epidural scar formation after lumbar spine surgery. Its positive effect concerning neural transmission at the level of the medulla was proven by electrophysiological tests in which the amplitude of components I and II of SSEP in the TachoComb group were significantly higher than in the control group.

Nodular pulmonary amyloidosis and Sjögren's syndrome in a patient treated with intermittent hemodialysis.

In the available literature, we have found the descriptions of 5 cases of nodular pulmonary amyloidosis associated with Sjögren's syndrome. In our practice, such a case has occurred in a patient with chronic renal failure. A 53-year-old woman underwent nephrological, rheumatological, and pulmonological examinations because of end-stage renal disease with a small cirrhotic kidney in renal ultrasound examination, pulmonary nodules, and xerophthalmia. Serological data revealed a slightly positive rheumatoid factor, antinuclear antibodies, anti-SS-A, anti-SS-B, and anti-RNP/Sm antibodies. Schirmer's test was positive on both sides and Sjögren's syndrome was recognized. Pulmonological examinations (the chest radiograph and CT scan, bronchofiberoscopy, culture of bronchial washings, bronchial biopsy, pleural effusion analysis, and a thick-needle biopsy) failed to determine the etiology of nodular changes in lungs. Immunofluorescence studies in the skin biopsy specimen showed IgM-positive staining. After 2 years of treatment with IHD, a toracoscopy was performed with enucleation of the nodules from the right lung. Histological examination showed massive deposits of amyloid, which allowed for a diagnosis of diffusive nodular pulmonary amyloidosis.

[Surgical treatment of spondylitic spine with transpedicular stabilisation]. / Leczenie operacyjne procesów zapalnych kregoslupa z zastosowaniem stabilizacji przeznasadowej.

BACKGROUND AND PURPOSE: Widespread use of antibiotics resulted in considerable reduction of spondylitis, but despite that progress there are some cases where conservative treatment has failed. In these patients surgical intervention should be carefully considered. Early surgery in patients with increasing neurological deficits causes the uncertainty which exists in relation to possible complications. Fewer doubts are associated with surgical treatment of late consequences (spine deformity, instability, etc). The goal of this study was to present indications for surgery, operative techniques of transpedicular stabilization and results of treatment of thoracic and lumbar spondylitis. MATERIAL AND METHODS: Clinical analysis includes 18 patients with spondylitis treated in the Department of Neurosurgery and Neurotraumatology University of Medical Sciences in Poznan between 1997 and 2004. There were 7 males (39%) and 11 females (61%) in the study group, and their mean age was 57+/-10 years (range 49-69). Spondylitis within thoracic segments was found in 15 (83%) cases and 3 (17%) patients had spondylitis within lumbar segments. Indications for early surgical intervention included increasing neurological symptoms, spine instability and failed conservative treatment. All patients underwent transpedicular stabilization following medulla and nerve root decompression. RESULTS: Non-specific inflammation was observed in 14 (78%) cases, and specific inflammation was discovered in 4 (22%) cases (bacteriological and histopathological findings). Very good and satisfactory results were achieved in 16 (89%) patients. Complications occurred in 4 (22%) patients. CONCLUSIONS: Spondylitic spine surgery limits inflammatory reaction, strengthens diagnosis, repairs vertebral column stability and improves neurological condition. Transpedicular stabilization is an effective method of achieving of internal spondylodesis.

[A case of a giant cavernoma located in the temporal region]. / Przypadek olbrzymiego naczyniaka jamistego plata skroniowego.

We present a case of a 21-year-old female with a giant cavernoma (5 cm x 3.1 cm x 3.6 cm) located in the left temporal region. Giant cavernomas are very rarely diagnosed. In our research we wanted to focus on the occurrence of these vascular malformations and diagnostic problems related to this matter.

Exploring viewing behavior data from whole slide images to predict correctness of students' answers during practical exams in oral pathology.

The way of viewing whole slide images (WSI) can be tracked and analyzed. In particular, it can be useful to learn how medical students view WSIs during exams and how their viewing behavior is correlated with correctness of the answers they give. We used software-based view path tracking method that enabled gathering data about viewing behavior of multiple simultaneous WSI users. This approach was implemented and applied during two practical exams in oral pathology in 2012 (88 students) and 2013 (91 students), which were based on questions with attached WSIs. Gathered data were visualized and analyzed in multiple ways. As a part of extended analysis, we tried to use machine learning approaches to predict correctness of students' answers based on how they viewed WSIs. We compared the results of analyses for years 2012 and 2013 - done for a single question, for student groups, and for a set of questions. The overall patterns were generally consistent across these 3 years. Moreover, viewing behavior data appeared to have certain potential for predicting answers' correctness and some outcomes of machine learning approaches were in the right direction. However, general prediction results were not satisfactory in terms of precision and recall. Our work confirmed that the view path tracking method is useful for discovering viewing behavior of students analyzing WSIs. It provided multiple useful insights in this area, and general results of our analyses were consistent across two exams. On the other hand, predicting answers' correctness appeared to be a difficult task - students' answers seem to be often unpredictable.

Association of galectin-1- but not galectin-3-dependent parameters with proliferation activity in human neuroblastomas and small cell lung carcinomas.

BACKGROUND: Galectins, a family of animal lectins binding beta-galactosides, are involved in growth regulation of diverse cell types in vitro, even harboring the potential to act as biphasic modulators with cell type selectivity. Owing to this capacity they might affect tumor growth when expression is adapted adequately. MATERIALS AND METHODS: To determine galectin-1-/-3- related features in routinely-fixed sections of two tumor types with poor prognosis (neuroblastoma and small cell lung carcinoma), immuno- and lectin histochemistry with specific antibodies and labeled galectins was performed. RESULTS: In comparison to previously studied tissue culture models, galectin-3 was frequently present, documenting occurrence of discrepancies between tumor models and clinical material for this protein. Cytoplasmic staining with galectin-1 and its antibody coincides with the proliferative activity of positive tumor cells determined by the MIB-1 monoclonal antibody. No statistical correlation was seen for galectin-3. CONCLUSION: These results encourage further cell biological studies to assess a regulatory role of galectin-1 on cell growth in vitro as a model for interfering with tumor proliferation by modulating expression of this type of endogenous effector(s).

Students' performance during practical examination on whole slide images using view path tracking.

BACKGROUND: Whole slide images (WSIs) used in medical education can provide new insights into how histological slides are viewed by students. We created software infrastructure which tracks viewed WSI areas, used it during a practical exam in oral pathology and analyzed collected data to discover students' viewing behavior. METHODS: A view path tracking solution, which requires no specialized equipment, has been implemented on a virtual microscopy software platform (WebMicroscope, Fimmic Ltd, Helsinki, Finland). Our method dynamically tracks view paths across the whole WSI area and all zoom levels, while collecting the viewing behavior data centrally from many simultaneous WSI users. We used this approach during the exam to track how all students (N = 88) viewed WSIs (50 per student) when answering exam questions (with no time limit). About 74,000 records with information about subsequently displayed WSI areas were saved in the central database. Gathered data was processed and analyzed in multiple ways. Generated images and animations showed view fields and paths marked on WSI thumbnails, either for a single student or multiple students answering the same question. A set of statistics was designed and implemented to automatically discover certain viewing patterns, especially for multiple students and WSIs. Calculated metrics included average magnification level on which a WSI was displayed, dispersion of view fields, total viewing time, total number of view fields and a measure depicting how much a student was focused on diagnostic areas of a slide. RESULTS: Generated visualizations allowed us to visually discover some characteristic viewing patterns for selected questions and students. Calculated measures confirmed certain observations and enabled generalization of some findings across many students or WSIs. In most questions selected for the analysis, students answering incorrectly tended to view the slides longer, go through more view fields, which were also more dispersed - all compared to students who answered the questions correctly. CONCLUSIONS: Designed and implemented view path tracking appeared to be a useful method of uncovering how students view WSIs during an exam in oral pathology. Proposed analysis methods, which include visualizations and automatically calculated statistics, were successfully used to discover viewing patterns. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/13000_2014_208.

Histopathologic patterns of nervous system tumors based on computer vision methods and whole slide imaging (WSI).

BACKGROUND: Making an automatic diagnosis based on virtual slides and whole slide imaging or even determining whether a case belongs to a single class, representing a specific disease, is a big challenge. In this work we focus on WHO Classification of Tumours of the Central Nervous System. We try to design a method which allows to automatically distinguish virtual slides which contain histopathologic patterns characteristic of glioblastoma--pseudopalisading necrosis and discriminate cases with neurinoma (schwannoma), which contain similar structures--palisading (Verocay bodies). METHODS: Our method is based on computer vision approaches like structural analysis and shape descriptors. We start with image segmentation in a virtual slide, find specific patterns and use a set of features which can describe pseudopalisading necrosis and distinguish it from palisades. Type of structures found in a slide decides about its classification. RESULTS: Described method is tested on a set of 49 virtual slides, captured using robotic microscope. Results show that 82% of glioblastoma cases and 90% of neurinoma cases were correctly identified by the proposed algorithm. CONCLUSION: Our method is a promising approach to automatic detection of nervous system tumors using virtual slides.

Whole slide imaging for educational purposes.

Digitized slides produced by whole slide image scanners can be easily shared over a network or by transferring image files to optical or other data storage devices. Navigation of digitized slides is interactive and intended to simulate viewing glass slides with a microscope (virtual microscopy). Image viewing software permits users to edit, annotate, analyze, and easily share whole slide images (WSI). As a result, WSI have begun to replace the traditional light microscope, offering a myriad of opportunities for education. This article focuses on current applications of WSI in education and proficiency testing. WSI has been successfully explored for graduate education (medical, dental, and veterinary schools), training of pathology residents, as an educational tool in allied pathology schools (e.g., cytotechnology), for virtual tracking and tutoring, tele-education (tele-conferencing), e-learning, virtual workshops, at tumor boards, with interactive publications, and on examinations. WSI supports flexible and cost-effective distant learning and augments problem-oriented teaching, competency evaluation, and proficiency testing. WSI viewed on touchscreen displays and with tablet technology are especially beneficial for education. Further investigation is necessary to develop superior WSI applications that better support education and to design viewing stations with ergonomic tools that improve the WSI-human interface and navigation of virtual slides. Studies to determine the impact of training pathologists without exposure to actual glass slides are also needed.

Quality evaluation of virtual slides using methods based on comparing common image areas.

BACKGROUND: There are many scanners of glass slides on the market now. Quality of digital images produced by them may be different and pathologists who examine virtual slides on a monitor may subjectively evaluate it. However, objective comparison of quality of digital slides captured by various devices requires assessment algorithms, which will be automatically executed. METHODS: In this work such an algorithm is proposed and implemented. It is dedicated for comparing quality of virtual slides which show the same glass slide captured by two or more scanners. In the first step this method looks for the largest corresponding areas in the slides. This task is realized by defining boundaries of tissues and providing the relative scale factor. Then, a certain number of smaller areas, which show the same fragments of both slides, is selected. The chosen fragments are analyzed using Gray Level Co-occurrence Matrix (GLCM). For GLCM matrices some of the Haralick features are calculated, like contrast or entropy. Basing on results for some sample images, features appropriate for quality assessment are chosen. Aggregation of values from all selected fragments allows to compare the quality of images captured by tested devices. RESULTS: Described method was tested on two sets of ten virtual slides, acquired by scanning the same set of ten glass slides by two different devices. First set was scanned and digitized using the robotic microscope Axioscope2 (Zeiss) equipped with AxioCam Hrc CCD camera. Second set was scanned by DeskScan (Zeiss) with standard equipment. Before analyzing captured virtual slides, images were stitched and converted using software which utilizes advances in aerial and satellite imaging.The results of the experiment show that calculated quality factors are higher for virtual slides acquired using first mentioned device (Axioscope2 with AxioCam). CONCLUSIONS: Results of the tests are consistent with opinion of the pathologists who assessed quality of virtual slides captured by these devices. This shows that the method has potential in automatic evaluation of virtual slides' quality.

Five years of experience teaching pathology to dental students using the WebMicroscope.

BACKGROUND: We describe development and evaluation of the user-friendly web based virtual microscopy - WebMicroscope for teaching and learning dental students basic and oral pathology. Traditional students microscopes were replaced by computer workstations. METHODS: The transition of the basic and oral pathology courses from light to virtual microscopy has been completed gradually over a five-year period. A pilot study was conducted in academic year 2005/2006 to estimate the feasibility of integrating virtual microscopy into a traditional light microscopy-based pathology course. The entire training set of glass slides was subsequently converted to virtual slides and placed on the WebMicroscope server. Giving access to fully digitized slides on the web with a browser and a viewer plug-in, the computer has become a perfect companion of the student. RESULTS: The study material consists now of over 400 fully digitized slides which covering 15 entities in basic and systemic pathology and 15 entities in oral pathology. Digitized slides are linked with still macro- and microscopic images, organized with clinical information into virtual cases and supplemented with text files, syllabus, PowerPoint presentations and animations on the web, serving additionally as material for individual studies. After their examinations, the students rated the use of the software, quality of the images, the ease of handling the images, and the effective use of virtual slides during the laboratory practicals. Responses were evaluated on a standardized scale. Because of the positive opinions and support from the students, the satisfaction surveys had shown a progressive improvement over the past 5 years. The WebMicroscope as a didactic tool for laboratory practicals was rated over 8 on a 1-10 scale for basic and systemic pathology and 9/10 for oral pathology especially as various students' suggestions were implemented. Overall, the quality of the images was rated as very good. CONCLUSIONS: An overwhelming majority of our students regarded a possibility of using virtual slides at their convenience as highly desirable. Our students and faculty consider the use of the virtual microscope for the study of basic as well as oral pathology as a significant improvement over the light microscope.

Genomic analysis of the 55 kDa subunit of DNA polymerase epsilon in human intracranial neoplasms.

BACKGROUND: Defects of some DNA polymerases have shown cancer associations, but there are only limited data on DNA polymerase (Pol) epsilon. MATERIALS AND METHODS: We examined 26 human brain neoplasm DNA samples and 8 control blood samples (from Poland) for possible mutations in the entire coding region of the 55 kDa small subunit of human DNA Pol epsilon gene using polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) analysis, and sequence analysis of DNA. RESULTS: One single base intronic transition in intron 14 was found. The AATT deletion previously found in some breast and colorectal tumors was not found in samples from brain neoplasms or controls, but it was found in 1/100 normal blood samples from South-West Finland. CONCLUSION: We found no evidence that potential mutations in the 55 kDa subunit of DNA Pol epsilon are a contributing factor in the development of the tested cases of human intracranial tumors.

A European network for virtual microscopy--design, implementation and evaluation of performance.

Web-based virtual microscopy has enabled new applications within pathology. Here, we introduce and evaluate a network of academic servers, designed to maximize image accessibility to users from all regions of Europe. Whole-slide imaging was utilized to digitize the entire slide set (n = 154) for the slide seminars of the 21st European Congress of Pathology. The virtual slides were mirrored to five academic servers across Europe using a novel propagation method. Functionality was implemented that automatically selects the fastest server connection in order to optimize the slide-viewing speed ( http://www.webmicroscope.net/ECP2007). Results show that during 6 months of monitoring the uptime of the network was 100%. The average viewing speed with the network was 3.1 Mbit/s, as compared to 1.9 Mbit/s using single servers. A good viewing speed (>2Mbit/s) was observed in 32 of 37 countries (86%), compared to 25 of 37 (68%) using single servers. Our study shows that implementing a virtual microscopy network spanning a large geographical area is technically feasible. By utilizing existing academic networks and cost-minimizing image compression, it is also economically feasible.

Genomic instability in pituitary adenomas.

Pituitary adenomas most commonly are identified as small, incidental microadenomas. They however may progress to macroadenoma forming intra and later suprasellar tumors which in about 1/3 of cases invade surrounding structures at the time of diagnosis. Mechanism of pituitary tumorigenesis remains still elusive. Because the value of karyotyping is limited by the technical problems related to cytogenetic methods, we studied the spectrum of chromosomal imbalances associated with pituitary adenoma using comparative genomic hybridization (CGH). Copy number aberrations on all 22 autosomes were evaluated by CGH using advanced computer software. In total, fifteen patients were included in the study of 9 non-invasive, 4 invasive and two recurrent adenomas. The mean age of the patients were 48 years ranging from 36 to 68 years. Five tumors showed hormonal activity. The histogram of all 15 cases representing the DNA imbalances as an incidence curve along each chromosome showed losses particularly for chromosomes 1p, 2q, 4, 5, 6, 11q, 12q, 13q and 18q as well as overrepresentation on 9q, 16p, 17p, 19, 20q. Functioning adenomas carried more imbalances than non-functioning, specifically deletions on chromosome 4 and 18q as well as overrepresentations of chromosomes 17 and 19. Invasive adenomas carried more overrepresentations at 1p34 than non-invasive tumors. Recurrent adenomas harbored more alterations than primary tumors, particularly DNA gains. The primary data is accessible at our CGH online tumor database at http://amba.charite.de/cgh. Reviewing the existing literature on the genetics of pituitary adenoma and discussing our results in this context, we hope that our study will contribute to the knowledge of this neoplasm.

Distinct methylation profiles of glioma subtypes.

Gliomas are tumors of the central nervous system with a wide spectrum of different tumor types. They range from pilocytic astrocytoma, with a generally good prognosis, to the extremely aggressive malignant glioblastoma. In addition to these 2 types of contrasting neoplasms, several other subtypes can be distinguished, each characterized by specific phenotypic, as well as genotypic features. Recently, the epigenotype, as evident from differentially methylated DNA loci, has been proposed to be useful as a further criterion to distinguish between tumor types. In our study, we screened 139 tissue samples, including 33 pilocytic astrocytomas, 46 astrocytomas of different grades, 7 oligoastrocytomas, 10 oligodendrogliomas, 10 glioblastoma multiforme samples and 33 control tissues, for methylation at CpG islands of 15 different gene loci. We used the semiquantitative high throughput method MethyLight to analyze a gene panel comprising ARF, CDKN2B, RB1, APC, CDH1, ESR1, GSTP1, TGFBR2, THBS1, TIMP3, PTGS2, CTNNB1, CALCA, MYOD1 and HIC1. Seven of these loci showed tumor specific methylation changes. We found tissue as well as grade specific methylation profiles. Interestingly, pilocytic astrocytomas showed no evidence of CpG island hypermethylation, but were significantly hypomethylated, relative to control tissues, at MYOD1. Our results show that glioma subtypes have characteristic methylation profiles and, with the exception of pilocytic astrocytomas, show both locus specific hyper- as well as hypomethylation.