MUTYH hotspot mutations in unselected colonoscopy patients.

AIM: Biallelic MutY human homologue (MUTYH) germline mutations predispose to recessively inherited adenomatous polyposis, designated MUTYH-associated polyposis (MAP), and colorectal cancer (CRC). The hotspot mutations p.Y179C and p.G396D account for the majority of pathogenic variants of MUTYH in Caucasians. Our aim was to evaluate the prevalence of MUTYH mutations in a prospective cohort of unselected patients with different colorectal diseases. METHOD: The hotspot mutations p.Y179C and p.G396D were genotyped in 352 consecutive patients undergoing colonoscopy at our tertiary referral centre. Exons 2-14 were sequenced in hotspot mutation carriers to exclude additional variants. RESULTS: Overall, we identified five heterozygous p.Y179C mutations and three heterozygous p.G396D mutations in seven hotspot mutation carriers (risk allele frequencies 0.7% and 0.4%, respectively). Two of these hotspot mutation carriers harboured a heterozygous p.Q338H variant, which is of uncertain clinical significance, on the other allele. Three individuals were biallelic MUTYH variant carriers (p.Y179C/p.G382D: typical MAP; p.Y179C/p.Q338H: atypical MAP with late onset and lower polyp burden; p.G382D/p.Q338H: inflammatory bowel disease), and four subjects were monoallelic mutation carriers. CONCLUSION: MUTYH-associated disease, and hence genetic counselling and MUTYH genetic testing, should be considered in the clinical routine of an endoscopy unit, but the wide range of phenotypes represents a challenge for patient identification. The clinical significance of p.Q338H should be evaluated in future case-control studies because compound heterozygotes for pathogenic mutations and p.Q338H may be at increased risk for mild polyposis or CRC. In addition, MUTYH should be assessed as a potential susceptibility gene for the development of colitis-associated CRC in future.

Characterization of the nuclear import of human MutLalpha.

DNA mismatch repair (MMR) is essential for the maintenance of replication fidelity. Its major task is to recognize mismatches as well as insertion/deletion loops of newly synthesized DNA strands. Although different players of human MMR have been identified, the regulation of essential steps of MMR is poorly understood. Because MMR is initiated in the nucleus, nuclear import might be a mechanism to regulate MMR. Nuclear targeting is accomplished by conserved signal sequences called nuclear localization signals (NLS), which represent clusters of positively charged amino acids (aa). hMLH1 contains two clusters of positively charged amino acids, which are candidate NLS sequences (aa 469-472 and 496-499), while hPMS2 contains one (aa 574-580). To study the effect of these clusters on nuclear import, NLS mutants of hMLH1 and hPMS2 were generated and expressed in 293T cells. The subcellular localization of the mutant constructs was monitored by confocal laser microscopy. We demonstrated that missense mutations of two signal sequences, one in hMLH1 and one in hPMS2, lead to impaired nuclear import, which was especially prominent for mutants of the hMLH1 residues K471 and R472; and hPMS2 residues K577 and R578.

Spontaneous fusion of 'os acetabuli' after triple pelvic osteotomy.

In acetabular dysplasia, an overloading of the acetabular rim can cause a stress fracture, creating an 'os acetabuli', or a lesion of the acetabular labrum. At puberty, the os acetabuli seems to be the epiphysis of the os pubis. We present the case of a 14-year-old girl with acetabular dysplasia and spontaneous fusion of an 'os acetabuli' after biomechanical correction by triple pelvic osteotomy. Our report supports the correctness of the biomechanical principle: reorientation of the acetabulum results in a better coverage of the femoral head, reduces the stress at the acetabular rim, shifts the os acetabuli out of the stress region, and may allow union of the bony fragment with the acetabulum.

Transient mismatch repair gene transfection for functional analysis of genetic hMLH1 and hMSH2 variants.

BACKGROUND: Germline mutations in the mismatch repair (MMR) genes hMLH1 and hMSH2 can cause hereditary non-polyposis colorectal cancer (HNPCC). However, the functional in vitro analysis of hMLH1 and hMSH2 mutations remains difficult. AIMS: To establish an in vitro method for the functional characterisation of hMLH1 and hMSH2 mutations. METHODS: hMLH1 and hMSH2 wild type (wt) genes and several mutated subclones were transiently transfected in mismatch repair deficient cell lines (HCT-116 and LOVO). Apoptosis, proliferation, and regulation of mRNA expression and protein expression of interacting proteins were analysed by Hoechst staining, AlamarBlue staining, real time polymerase chain reaction, and western blotting, respectively. RESULTS: The protein expression of hMLH1 and hMSH2 mutants was significantly decreased after transfection compared with wild type transfections. The hMLH1 and hMSH2 interacting proteins hPMS2 and hMSH6 became detectable only after transfection of the respective wild type genes. In parallel, hMSH6 mRNA levels were increased in hMSH2 wt transfected cells. However, hPMS2 mRNA levels were independent of the mutation status of its interacting partner hMLH1, indicating a post-transcriptional regulating pathway. In the hMLH1 deficient HCT-116 cell line apoptosis was not affected by transfection of any mismatch repair gene, whereas complementation of hMSH2 deficiency in LOVO cells increased apoptosis. Conversely, proliferative activity of HCT-116 was decreased by complementation with hMLH1wt and unaffected in hMSH2 deficient LOVO cells. CONCLUSION: These data show that the cellular role of the MMR genes and its mutations are assessable in a simple transient transfection system and show the influence of MMR gene regulation on major cell growth regulating mechanisms. This method is applicable for the functional definition of mutations in hMLH1 and hMSH2 genes observed in patients with suspected HNPCC.

Bethesda guidelines: relation to microsatellite instability and MLH1 promoter methylation in patients with colorectal cancer.

BACKGROUND: Microsatellite instability is a hallmark of mismatch repair deficiency in hereditary nonpolyposis colorectal cancer and results from mutations in the mismatch repair genes MLH1 or MSH2 or from gene inactivation associated with DNA methylation. The Bethesda guidelines were established to identify patients with colorectal cancer who should be tested for microsatellite instability. OBJECTIVE: To assess the Bethesda guidelines for detection of microsatellite instability and to determine the role of MLH1 promoter methylation in colorectal cancer. DESIGN: Prospective cohort study. SETTING: Tertiary care referral center in Frankfurt, Germany. PATIENTS: 125 consecutive patients with colorectal cancer. MEASUREMENTS: Patients were assessed according to the Bethesda guidelines, and tumor specimens were analyzed for microsatellite instability. Patients with microsatellite instability were tested for MLH1 promoter methylation and MLH1 and MSH2 germline mutations. RESULTS: Microsatellite instability was detected in 17 of 58 patients who fulfilled and 5 of 67 patients who did not fulfill criteria of the Bethesda guidelines. In 11 of 17 patients with microsatellite instability who fulfilled Bethesda guidelines, an MLH1 (n = 3), MSH2 (n = 7), or combined MLH1 and MSH2 (n = 1) mutation was found. Among the patients with microsatellite instability who did not fulfill Bethesda guidelines, no mutations were observed; MLH1 promoter methylation was observed in 6 of 11 patients with an MLH1 or MSH2 mutation and 5 of 11 patients without an MLH1 or MSH2 mutation. CONCLUSIONS: The Bethesda guidelines are useful for selecting patients for microsatellite instability testing. MLH1 and MSH2 testing should be recommended in all patients with colorectal cancer and microsatellite instability who fulfill at least one Bethesda criterion. MLH1 promoter methylation may accompany rather than initiate carcinogenesis in patients with colorectal cancer who have mismatch repair gene defects.

Magnetic resonance arthrography of the acetabular labrum: value of radial reconstructions.

The intention was to compare the sensitivity and specificity of radial planes with oblique coronal and oblique axial planes for the detection of labral abnormalities of the acetabulum on magnetic resonance arthrography (MRa). Twenty cadaveric hip joints were examined by radiography and MRa. For MRa, 15 ml of a solution of iodinated contrast material and gadolinium diethylene triamine tetra-acetic acid (Gd-DTPA; 100:1) were injected under fluoroscopic guidance. MRI was performed on a 1.5-T MR scanner with a fat-suppressed 3D-FLASH sequence (TR/TE/flip-angle 42 ms/10 ms/40 degrees; field of view 16 cm, matrix 256 x 256, section thickness 1.5 mm, pixel size 0.7 x 0.7 mm). Multiplanar image reconstructions were done perpendicular to the acetabulum in oblique coronal and oblique axial planes and in radial planes. Macroscopic and histopathologic examination of the labral specimens was performed. Labral lesions were found in 15/20 hips (75%) on pathologic examination. Six hips demonstrated labral degeneration. The labrum was partially detached in 7 hips and completely detached in 2 hips. A flap-like labrum was found in 2 cases, 1 with partial detachment of the labrum and 1 with a degenerated labrum. Using oblique coronal and oblique axial reconstructions, pathologic findings were confirmed by MRa in 9/15 specimens (sensitivity 60%). There were no false-positive findings (specificity 100%, accuracy 70%). Also, 3/6 labral degenerations without detachment, 4/7 partial detachments, and 2/2 complete detachments were correctly diagnosed. Two flap-like labra were not recognized. With radial reconstructions, pathologic findings were correctly confirmed in 12/15 specimens (sensitivity 80%) without false-positive findings (specificity 100%, accuracy 85%). Also, 3/6 labral degenerations without detachment, 6/7 partial detachments, 2/2 complete detachments, and 1/2 flap-like labra were correctly diagnosed. MRa of the acetabular labrum using radial reconstructions is well suited to delineate the acetabular labrum and to diagnose labral detachments. Radial reconstructions allow for perpendicular display of the whole acetabular circumference and are more accurate for the diagnosis of acetabular labral lesions, when compared with oblique coronal and oblique axial reconstructions.

Activation of a cryptic splice site of PTEN and loss of heterozygosity in benign skin lesions in Cowden disease.

Cowden disease is an autosomal dominant syndrome characterized by facial trichilemmomas, acral keratoses, papillomatous papules, mucosal lesions, and an increased risk for breast and nonmedullary thyroid cancer. Here, we describe a novel PTEN splicing site mutation in a family with classical Cowden disease and we studied benign skin lesions typical for Cowden disease for loss of heterozygosity. We found a PTEN IVS2 + 1G > Alpha 5'-splicing acceptor mutation resulting in activation of a cryptic splice site. Activation of this cryptic splice site is predicted to result in a frameshift with a premature stop codon, thus disrupting the phosphatase core motif of PTEN. Loss of heterozygosity analysis of two trichilemmomas, one fibroma, and three acanthomas of the index patient demonstrated loss of heterozygosity at the PTEN locus in four of these lesions. In conclusion, our data demonstrate that a PTEN splicing site mutation causes activation of a cryptic splice site, which results in aberrant transcripts.

Magnetic resonance arthrography of the acetabular labrum. Macroscopic and histological correlation in 20 cadavers.

We studied the sensitivity and specificity of magnetic resonance arthrography (MRa) for the diagnosis of lesions of the acetabular labrum in 20 cadaver hips. The MRa results were compared with macroscopic and histological findings. We found that the labrum could be satisfactorily delineated by MRa and that large detachments could be identified satisfactorily. The diagnosis of small detachments and degeneration of the labrum was less reliable.

[Polyethylene content and cell size of foreign body giant cells in aseptic hip prosthesis loosening--a histomorphometric study]. / Polyethylenbeladung und Zellgrösse von Fremdkörperriesenzellen bei der aseptischen Hüftendoprothesenlockerung--Eine histomorphometrische Untersuchung.

INTRODUCTION: In aseptic loosening of cementless hip arthroplasty polyethylene particles are phagocytized by macrophages and foreign-body giant cells. This initiates an osteolytic cascade. In this study, the authors investigated if there are correlations between the size of foreign-body giant cells and the polyethylene loading and number of intracellular particles as determined by light microscopy. MATERIAL AND METHOD: Histological specimens were processed from tissues retrieved during revision surgery of 7 cases of cementless hip arthroplasty which had become aseptically loose. The specimens were analyzed by histolomorphology and histomorphometry. The cell size, polyethylene loading and intracellular particle number of 111 foreign-body giant cells were determined. A regression analysis was performed to investigate if there was a correlation between these variables. RESULTS: The mean cell size was 1417 +/- 487 micron 2, the mean polyethylene loading was 49 +/- 42 micron 2 and the mean intracellular particle number was 10.4 +/- 5.4. The cell size correlated with both the intracellular particle number (r = 0.25) and the polyethylene loading (r = 0.39). CONCLUSIONS: In the cellular reaction to polyethylene particles in aseptic loosening of cementless total hip arthroplasty, the size of foreign-body giant cells correlates with intracellular polyethylene loading and intracellular polyethylene particle number as determined by light microscopy. The presence of large foreign body giant cells might be associated with a high particle load of the tissue and a high osteolytic activity.

[MR arthrography of the labrum acetabulare--a radiological-anatomical correlation in 20 cadaveric hips]. / MR-Arthrographie des Labrum acetabulare--radiologisch-anatomische Korrelation an 20 Leichenhüften.

PURPOSE: To investigate frequency of acetabular labral lesions in elderly hip joints, and to determine sensitivity and specificity of MR arthrography (MRa) for the detection of these abnormalities. MATERIALS AND METHODS: Twenty cadaveric hip joints were examined by MRa. For MRa, 15 ml of a solution of iodinated contrast solution (Solutrast 300) and Gd-DTPA (100:1) were injected under fluoroscopic guidance. MR imaging was performed on a 1.5 T MR scanner (Vision, Siemens; FOV 16 cm, matrix 256 x 256, fat-suppressed 3D-FLASH). Multiplanar image reconstructions were done perpendicular to the acetabulum in the oblique-coronal, oblique-axial, and radial planes. The labral specimens were examined macroscopically. RESULTS: In 12/20 hips (60%), a labral lesion was found on pathologic examination. In 7 specimens, the labrum was partially or completely detached in the weight-bearing superior region. One flap-like variant of the labrum was seen; in 4 hip joints, the labrum was degenerated (one cystic degeneration). Pathologic findings were confirmed by MRa in 8/12 specimens (sensitivity 67%). All degenerated labra were correctly diagnosed on MRa. Three small labral detachments and the flap-like variant were misinterpreted as being normal. There were no false positive findings (specificity 100%). The accuracy was 80%. Labral lesions were seen in 6/8 and in 6/12 of hips with and without osteoarthritis, respectively. CONCLUSION: MRa is well suited to delineate the acetabular labrum and to diagnose labral abnormalities. Detection of small labral detachments and anatomic variants is difficult and requires some experience. Labral lesions are correlated to osteoarthritis of the hip, but may be frequently seen in the elderly without underlying osteoarthritis.

Occipitocervical fusion in patients with rheumatoid arthritis.

Instability and deformity of the cervical spine caused by rheumatoid arthritis is a well known entity. Operative intervention is indicated for patients with progressive deformity and when pain is resistant to conservative treatment. In a series of 39 patients who underwent posterior occipitocervical fusion with a Y plate, 22 patients were observed clinically and radiographically at average 41.5 months after surgery. In 35 of the 39 patients the main indication for surgery was pain, and in 30 of the 39 patients additional neurologic deficit (radiculopathy or myelopathy) was present. Thirty-one of the 39 patients had atlantoaxial instability. The atlantoaxial instability was associated with cranial migration of the dens in 19 patients. According to the classification of Conaty and Mongan 77.3% patients had satisfactory results and 22.7% had unsatisfactory results. Of the 30 patients with neurologic deficit, nine patients had a significant improvement. No patient had a worse result after surgery. Solid fusion was seen in all 22 patients at followup. Seven patients experienced complications directly related to the surgical procedure. Posterior fixation combined with anterior decompression in the presence of spinal stenosis represents a useful and safe method to treat instability and deformity caused by rheumatoid arthritis. Early surgical procedures may reduce the complication rate.

[Dorsal occipitocervical fusion--indications and results]. / Die dorsale occipitozervikale Fusion--Indikationen und Resultate.

STUDY DESIGN: This report will relate to our experience with the occipitocervical fusion with the y-plate, which we used in the treatment of 53 patients. METHODS: 39 of the patients had rheumatoid arthritis. Other indications demanding surgery were posttraumatic conditions, degenerative and congenital lesions, osteomyelitis, tumor and psoriasis arthritis. Before surgery, all patients suffered from pain in the neck and/or in the back of the head. On a linear scale from 0 to 10, the pain was rated as 8.0 in average (range 4 to 10). 33 patients had an instability of the atlantoaxial region and 26 patients a basilar invagination of the odontoid. A cervical myelopathy was found in 23 cases. One surgeon fused the occiput to C2 in 30 cases, to C3-C5 in 14 cases and to C7-T2 in 9 cases. In 17 patients a resection of the odontoid had to be performed before fusion to adequately decompress the spinal cord. RESULTS: 31 of the patients could be controlled with a follow-up of at average 45.9 months. 16 patients had died. At the time of follow-up, the pain was rated as 2.1 in average (range 0 to 8). The myelopathy cleared up in all cases but 3.9 patients required further operations on the cervical spine. 5 patients developed an instability at the level(s) below the fusion and an enlargement of the fusion to these levels had been performed. The fusion rate was 98.1%. The results were satisfying in 25 (80.6%) and not satisfying in 5 patients (19.4%). CONCLUSIONS: These results show the effectiveness of the occipitocervical fusion with the y-plate in a wide range of applications. It offers advantages above other techniques.

[Occipitocervical fusion in chronic polyarthritis]. / Die okzipitozervikale Fusion bei chronischer Polyarthritis.

This report will relate our experience with the y-plate, which we used in the treatment of 39 patients (32 women and 7 men) with a mean age of 62.6 years (range 47 to 79 years) between 1987 and 1994. All patients had rheumatoid arthritis. Occipitocervical fusion was indicated by instabilities within the occipitocervical region and in cases with additional basilar invagination and/or after transoral dens resection for decompression of the spinal cord. The length of the fusion depended on the pathological changes of the subaxial cervical spine. Before surgery, 35 patients suffered from strong and 4 patients from moderate pain in the neck and/or the back of the head. On a linear scale from 0 to 10, the pain was rated as 8:1 on average (range 4 to 10). 31 patients had an instability of the atlantoaxial region and 19 patients a basilar invagination of the odontoid. A cervical myelopathy was found in 20 cases. One surgeon fused the occiput to C2 in 22 cases, to C3-C5 in 8 cases and to C7-T2 in 9 cases. An important factor in this operation is the integration of atlantoaxial screws in order to resist the translational dislocation of C1/C2. In 13 patients a resection of the odontoid had to be performed to adequately decompress the spinal cord. A reduction of C2 without dens resection was performed in cases with reducible instability C1/2. 32 of the patients could be controlled with a minimum follow-up of 12 months (average 32.2 months, range 12 to 66 months). Out of the other 7, 6 patients had died. At the time of follow-up, the pain was rated as 2.3 on average (range 0 to 10). A myelopathy was present in 2 cases. Six patients required further operations on the cervical spine; 4 patients developed an instability at the level(s) below the fusion and an enlargement of the fusion to these levels has been performed. The fusion rate was 96.9%, despite breakage of the implant in 3 and a screw loosening in 2 patients. According to the criteria of Conaty, the result was satisfying in 25 (75%) and not satisfying in 8 patients (25%). These results show the effectiveness of the occipitocervical fusion with the y-plate in rheumatoid arthritis. A transoral dens resection is only indicated in cases with basilar invagination causing a compression of the spinal cord or in such cases where a compression caused by the dens or the retrodental pannus formation cannot be treated by a reduction of the second cervical vertebra alone.

Anterior lumbar vertebral translation following translaminar screw fixation. A report of five cases.

Translaminar screw fixation is a good procedure for posterolateral spinal fusion of one or two motion segments. Anterior translation of the upper fused vertebra occurred in 5 of our patients following this procedure. A further fusion with pedicle screw fixation was needed in 2 of them. This complication has been previously reported after decompression operations on the lumbar spine. Translaminar screwing produces slight posterior distraction which pushes forward the upper vertebra. This procedure should be avoided in cases with even minimal anterior translation before operation. Translaminar screw fixation is an ideal technique for fusion of a degenerated segment when the upper vertebra has slipped posteriorly.

["Recurrent" epiphysiolysis capitis femoris--need for simultaneous stabilization of both hip joints]. / "Rezidivierende" Epiphysiolysis capitis femoris--die Notwendigkeit der simultanen Stabilisierung beider Hüftgelenke.

In the operative therapy of slipped capital femoral epiphysis, a controversial discussion exists about the sense of a simultaneous stabilization of both epiphyseal plates and about the correct time of removing the implants. We show the epikrisis of two patients to demonstrate our therapy regimen: in the first case the implants were removed too early. The consequence was a further slipping of the femoral epiphysis consecutive on both sides and the necessity of further surgery. In the other case we observed an outgrowing of the implant out of the epiphyseal plate by use of unthreaded wires. The growth plate was not closed. The femoral epiphysis was stabilized again, because an outgrowing compares to an implant removal at a too early stage. These two cases show our point of view. We recommend to stabilize both sides simultaneously with threaded wires and not to remove the implants before closure of the epiphyseal plate. Furthermore, we hold it advisable to perform a restabilization, when the implants grow out of the unclosed epiphyseal plate during adolescence.

[Micro-technological anterior discectomy without fusion in cervical disk displacement with radicular symptoms]. / Die mikrotechnische vordere Diskektomie ohne Fusion beim zervikalen Bandscheibenvorfall mit radikulären Beschwerden.

Only cervical disc herniation that provokes root compression unresponsive to conservative treatment should be selected for operation. The operative technique must allow adequate removal of the disc and relieve any root pressure without distressing the patient. Operative decompression of the root can be attained via laminectomy and arthrotomy or by way of an anterior approach. With this second procedure, the ventral discectomy can be done in conjunction with an interbody fusion. We report our experience with 216 patients who underwent anterior cervical microsurgical discectomy without fusion between 1980 and 1944. All these patients were suffering from compressive cervical radiculopathy caused by disc displacement without significant degenerative deformation of the motion segment and without manifest segmental instability. The follow-up ranged between 6 and 185 months (average 71 months) in the 175 patients in whom it was possible. Only in 7 patients (4%) is the result unsatisfactory; 79 patients (45%) are completely free of symptoms 99; (56.6%) are very satisfied and 45 (25.7%) satisfied with the result of the procedure.

Surgical treatment of degenerative spondylolisthesis in the lumbar spine: no reposition without prior decompression.

In the present note, the cause of root compression in cases with degenerative lumbar spondylolisthesis is discussed in order to clarify the best method of surgery and to avoid iatrogenic damage of the cauda equina if a transpedicular fusion is going to be performed. The root compression is due to osteophytes growing out from the upper articular facets and from the upper ridge of the lamina of the lower vertebra. Therefore, the repositioning of the anteriorly displaced vertebra by the instrumentation could lead to the squeezing of the cauda equina between the withdrawn vertebral body and the degeneratively enlarged and hypertrophic facet joints of the lower vertebra. The reposition (even if incomplete) must always be preceded by decompression on both sides, even if radicular complaints are unilateral.