Cognitive conditioning: imagined stimulus contiguity and the third interval conditional GSR.
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Conditioning, Classical↗
Biomedical subjects
M Werner
Publications and source records attributed to M Werner.
Iliopsoas abscess: a report of 24 patients diagnosed by CT.
BACKGROUND: We wanted to define the role of computed tomography (CT) in the diagnosis, etiology, and treatment of iliopsoas abscess. METHODS: Twenty-four patients (18 men, six women; age range = 17-86 years) with iliopsoas abscesses diagnosed over 8 years were retrospectively reviewed. All presented with fever and elevated white blood cell counts. Twenty-one had abdominal, flank or pelvic pain and nine had specific psoas signs suggesting the diagnosis. RESULTS: Seventeen of the abscesses were right-sided. Twenty were regarded as secondary to various underlying causes that were clearly demonstrated on CT and related to gastrointestinal (n = 12), skeletal (n = 5), or urinary tract (n = 3) diseases. All patients received appropriate antibiotic treatment. Thirteen also had their abscesses drained and eight had definitive surgical procedures. CONCLUSION: CT is an effective imaging technique for diagnosing iliopsoas abscess, even when classic clinical signs are absent. Treatment by percutaneous drainage under CT guidance is another advantage. When a psoas abscess is a complication of Crohn's disease, resection of the affected bowel segment is recommended in addition to drainage because drainage alone even in conjunction with appropriate medical therapy is usually not effective.
Adult↗
Target and signal amplification: approaches to increase the sensitivity of in situ hybridization.
In situ hybridization (ISH) has proven to be a very important molecular tool in research and diagnosis. However, its applicability can be limited by its restricted detection sensitivity. During the last few years, several strategies have been developed to improve the threshold levels for ISH detection by amplification of either target nucleic acid sequences prior to ISH or the detection signals after hybridization procedures. In this overview, we outline and analyze the principles, applications, and limitations of in situ polymerase chain reaction, in situ self-sustained sequence replication, primed in situ labeling (PRINS), and in situ transcription as examples of target amplification methods, and catalyzed reporter deposition using biotinylated tyramine as an approach to signal amplification in ISH. We also provide a detailed, 1-day protocol for non-radioactive oligonucleotide ISH including signal amplification, which is suitable for diagnostic purposes. Furthermore, future directions of ISH including combined strategies of target and signal amplification as well as automation are discussed.
Forecasting↗
Interphase cytogenetics in pathology: principles, methods, and applications of fluorescence in situ hybridization (FISH).
Characteristic chromosome aberrations have been identified in various tumors. Fluorescence in situ hybridization (FISH) using specific probes that are generated by vector cloning or in vitro amplification and labeled with fluorescent dyes allow for the detection of these genetic changes in interphase cells. This technique, that is also referred to as "interphase cytogenetics", can be performed in cytological preparations as well as in sections of routinely formaldehyde-fixed and paraffin-embedded tissue. In cancer research and diagnostics, interphase cytogenetics by FISH is used to detect numerical chromosome changes and structural aberrations, e.g., translocations, deletions, or amplifications. In this technical overview, we explain the principles of the FISH method and provide protocols for FISH in cytological preparations and paraffin sections. Moreover, possible applications of FISH are discussed.
Cytological Techniques↗
Comparative genomic hybridisation for the analysis of chromosomal imbalances in solid tumours and haematological malignancies.
Comparative genomic hybridisation (CGH) is based on a two-colour, competitive fluorescence in situ hybridisation of differentially labelled tumour and reference DNA to normal metaphase chromosomes. This new technology has made a great impact in molecular tumour pathology due to its possible application to archival specimens and the ability to create copy number karyotypes throughout the whole genome from very small amounts of DNA. If chromosomal imbalances can be correlated with a etiological and clinical features of tumours, CGH could be able to provide new prognostic and diagnostic criteria. CGH findings further provide starting points for the molecular genetic characterisation of altered chromosomal regions harbouring yet unidentified genes involved in tumorigenesis and tumour progression. An overview of the results of published CGH studies on solid tumours and haematological malignancies is presented. Methodological limitations of the CGH technology are reported, as well as future developments which will improve its use in routine analysis.
Chromosome Aberrations↗
Single- and double-color oligonucleotide primed in situ labeling (PRINS): applications in pathology.
Molecular cytogenetics is mostly performed by fluorescence in situ hybridization using long DNA probes that are generated by vector cloning. Oligonucleotide primed in situ labeling (PRINS) is a recent method that has been established for the detection of the centromeric or telomeric region in metaphase chromosomes. In this overview, we demonstrate the possible applications of PRINS and provide elaborated protocols for its use in intact interphase cells of routine cytological preparations, e.g., cell smears, touch preparations, and cytospins of non-neoplastic and neoplastic tissues. Moreover, the various modifications of the PRINS method, such as multi-color PRINS for targeting different chromosomes within one cell or the enzymatic detection of the PRINS product instead of the more commonly used fluorochromes, are discussed.
Centromere↗
The sulfoxidation of the hexachlorobutadiene metabolite N-acetyl-S-(1,2,3,4,4-pentachlorobutadienyl)-L-cysteine is catalyzed by human cytochrome P450 3A enzymes.
The sulfoxidation of the mercapturic acid N-acetyl-S-(1,2,3,4,4-pentachlorobuta-1,3-dienyl)-L-cysteine (N-Ac-PCBC), a urinary metabolite of the renal toxin hexachlorobutadiene (HCBD), was studied in human liver microsomes and with purified cDNA expressed human liver cytochrome P450 (P450) enzymes. N-Acetyl-S-(1,2,3,4,4-pentachlorobuta-1,3-dienyl)-L-cysteine sulfoxide (N-Ac-PCBC SO) is a major urinary metabolite of HCBD in male rats; only liver microsomes from male rats catalyze the sulfoxidation of N-Ac-PCBC. Our results presented here show that human liver microsomes from both male and female donors are capable of oxidizing N-Ac-PCBC to the corresponding sulfoxide diastereomers. The correlation of N-Ac-PCBC sulfoxidation with the rates of oxidation of P450 enzyme specific substrates suggests that only P450 3A enzymes oxidize N-Ac-PCBC. Moreover, only gestodene and troleandomycin, two selective inhibitors for P450 of the 3A family, significantly reduced the rates of N-Ac-PCBC sulfoxidation. No reduction in sulfoxidation rates was observed with inhibitors for other P450 enzymes, i.e., diethyldithiocarbamate, 4-methylpyrazole, 7,8-benzoflavone, or sulfaphenazole. Incubations of N-Ac-PCBC with purified and reconstituted recombinant P450s 1A2, 2E1, 3A4, and 3A5 resulted in sulfoxide formation only with P450s 3A4 and 3A5. In summary, these results indicate that P450 from the 3A family may sulfoxidize N-Ac-PCBC. Since these P450 enzymes account for a major fraction of the P450 in human liver and are also present in human kidney, the sulfoxidation reaction may also be expected to occur in humans exposed to HCBD.
Acetylcysteine↗
Sulfoxidation of mercapturic acids derived from tri- and tetrachloroethene by cytochromes P450 3A: a bioactivation reaction in addition to deacetylation and cysteine conjugate beta-lyase mediated cleavage.
In the present study we investigated the formation of sulfoxides from N-acetyl-S-(1,2,2-trichlorovinyl)-L-cysteine (N-Ac-TCVC), N-acetyl-S-(1,2-dichlorovinyl)-L-cysteine (N-Ac-1,2-DCVC), and N-acetyl-S-(2,2-dichlorovinyl)-L-cysteine (N-Ac-2,2-DCVC), which are formed in the glutathione dependent bioactivation of tri- and tetrachloroethene. The first aim was to elucidate the enzymes involved in these oxidation reactions. N-Ac-TCVC, N-Ac-1,2-DCVC, and N-Ac-2,2-DCVC are oxidized to the corresponding sulfoxides mainly, if not exclusively, by cytochrome P450 enzymes in liver microsomes of untreated male rats, since no role for the flavin-containing monooxygenase (FMO) could be demonstrated by heat inactivation experiments and by the use of n-octylamine. The sulfoxidation rates were increased when using liver microsomes of phenobarbital and dexamethasone pretreated male rats as well as liver microsomes of dexamethasone pretreated female rats, while no sulfoxide formation was observed in liver microsomes of untreated female rats, suggesting an involvement of cytochrome P450 3A. Also, troleandomycin, a specific chemical inhibitor for cytochrome P450 3A, drastically reduced sulfoxidation rates. The observed rates of sulfoxidation also correlated well with the rates of oxidation of testosterone at the 6-beta-position, a specific marker for P450 3A activity. The second aim of this study was to compare the cytotoxicity of the sulfoxides with the cytotoxicity of the corresponding mercapturic acids in isolated rat renal epithelial cells. Both mercapturic acids and the corresponding sulfoxides were cytotoxic. Cytotoxicity of the mercapturic acids could be blocked by (aminooxy)acetic acid (AOAA), an inhibitor of cysteine conjugate beta-lyase, while the cytotoxicity of the sulfoxides was not influenced by this treatment. Moreover, the sulfoxides were significantly more cytotoxic than the corresponding mercapturic acids at equimolar doses. The results show that mercapturic acids derived from TRI and PER are oxidized to sulfoxides by microsomal monooxygenases from rat liver. The cytotoxicity of the produced sulfoxides could not be reduced by AOAA, consistent with a role of the sulfoxides as direct acting electrophiles (i.e., Michael acceptor substrates).
Acetylation↗
Is urinary drainage necessary during continuous epidural analgesia after colonic resection?
BACKGROUND AND OBJECTIVES: Postoperative urinary retention may occur in between 10% and 60% of patients after major surgery. Continuous lumbar epidural analgesia, in contrast to thoracic epidural analgesia, may inhibit urinary bladder function. Postoperative urinary drainage has been common in patients with continuous epidural analgesia, despite the lack of scientific evidence for its indication after thoracic epidural analgesia. This study describes 100 patients who underwent elective colonic resection with 48 hours of continuous thoracic epidural analgesia and only 24 hours of urinary drainage. METHODS: This is a prospective, uncontrolled study with well-defined general anesthesia, postoperative analgesia, and nursing care programs in patients with a planned 2-day hospital stay, urinary catheter removal on the first postoperative morning, and epidural catheter removal on the second postoperative morning. Follow-up in the outpatient clinic was on days 8 and 30. RESULTS: Nine patients needed bladder recatheterization, 8 as a single procedure and 1 patient a second recatheterization with removal on day 7. This patient had urinary infection on day 10 and was readmitted for 5 days because of urosepsis and, subsequently, for cystitis and left-sided epididymitis. Three patients had uncomplicated urinary infection. No patients had urological complaints at 30 days follow-up (95% confidence limit, 0% to 3.6%). CONCLUSION: The low incidence of urinary retention (9%) and urinary infection (4%) suggests that routine bladder catheterization beyond postoperative day 1 may not be necessary in patients with ongoing continuous low-dose thoracic epidural analgesia.
Adult↗
[Tumors of the footbones- a report from the Hamburg Bone Tumor Registry].
Aim of the present study was a systematic examination of the tumors of the foot registered in the Hamburg Bone Tumor Registry regarding age, sex, location, diagnosis and morphological characteristics. The registry files and histological specimens of 367 tumors and tumor-like lesions of the foot were reviewed. Males were more frequently affected (ratio 4 : 3). Some 20.4 % of all mass forming tumors were malignant. Cartilage tumors (146 cases), cystic (132 cases) and osteoblastic lesions (33 cases) were most common. Several tumors showed a strong predilection for certain parts of the foot. In the calcaneus a solitary bone cyst was the most common lesion. Lesions in the talus and metatarsals with a similar morphology mostly represented intraosseous ganglion cysts. Other tumors in the calcaneus were mainly cartilaginous. Besides chondroblastoma, chondrosarcoma was regularly diagnosed. Chondroma was extremely rare at this site. Contrary to that chondroma was regularly found at the metatarsal and phalangeal bones, it was 4 times more common than chondrosarcoma. An even distribution showed aneurysmal bone cysts and osteoidosteoma. Very rare lesions were fibrous dysplasia, eosinophilic granuloma and intraosseous lipoma. The knowledge of the distribution of tumors in the foot is important for an exact differential diagnosis and thereby for biopsy planning, diagnosis and therapy.
Adult↗
[Calcaneal cyst: a classical simple bone cyst?].
INTRODUCTION: In the calcaneus differentiation between a solitary cyst and intraosseus lipoma is difficult. Radiologists frequently diagnose an intraosseus lipoma, whereas histology shows the classical signs of a solitary bone cyst. We present 12 cases of a solitary cyst of the calcaneus. MATERIAL AND METHODS: Between 1993 and 2001 we operated on 12 patients (8 men, 4 women, median age 28 +/- 14 years) with calcaneal cysts. 9 patients received a curretage and an autologous, one patient a homologous, one patient a mixed autologous-homologous bone-grafting and one patient only a curretage (due to the small diameter of the cyst) of their cysts. RESULTS: No case revealed the histological picture of a fatty formation, which appears to be typical for an intraosseus lipoma. All of the microscopic findings resembled the characteristics seen in cysts of the long bones. A pathological fracture has not been observed. CONCLUSION: The histologically confirmed calcaneal cysts showed the radiological signs that are supposed to be typical for an intraosseous lipoma. It cannot be decided whether the histologically diagnosed calcaneal lipomas described by others can be interpreted as fatty degeneration of a calcaneal cyst, or whether fatty areas of the bone marrow have given a wrong impression. Since the literature only describes single isolated cases of a pathological fracture of the calcaneal cyst or lipoma, asymptomatic patients should be treated non-operatively.
Adolescent↗
[Extrathoracal pulmonary sequestration in the differential diagnosis of paravertebral tumors in the neonate].
BACKGROUND: Paravertebral masses of the fetus are often detected during routine prenatal ultrasonography. The most common differential diagnoses of these tumors are neuroblastoma, adrenal hemorrhage, schwannoma and germ cell tumors. CASE REPORT: We report on a mature male newborn, who was diagnosed antenatally at 23 + 3 weeks of gestation with a tumor in the left paravertebral region. After birth the child was transferred to a neonatal unit. Tumor markers like urinary catecholamines were within normal limits, neuron-specific enolase was slightly elevated. MRI as well as ultrasonography confirmed a 3.2 x 2.2 x 1.6 cm large smoothed edged tumor in the left paravertebral region at the level of T10-T12. An open biopsy was performed, and the tumor which was located below the diaphragma was subtotally resected. Histopathology showed an extralobar pulmonary sequestration. Surgery as well as postoperative course was uneventful. CONCLUSIONS: Extralobar pulmonary sequestrations represent rare congenital anomalies, which are usually asymptomatic. Clear differentiation between tumor and pulmonary sequestration is seldom possible despite high resolution imaging studies. Hence, a biopsy procedure should be done for diagnosis of paravertebral masses.
Biopsy↗
[Neoadjuvant therapy for localized osteosarcoma of extremities. Results from the Cooperative osteosarcoma study group COSS of 925 patients].
BACKGROUND: Owing to twenty years of multicentric interdisciplinary cooperation, the COSS group has been able to collect data on a large group of osteosarcoma patients treated by neoadjuvant therapy. This paper reviews results achieved in patients with localized extremity tumors. PATIENTS AND METHODS INCLUSION CRITERIA: Registration into a completed neoadjuvant COSS-Study. Histologically confirmed, primary, localized, high-grade, central osteosarcoma of an extremity; age < 40 years; no pretreatment; interval diagnosis to chemotherapy < or = 3 weeks; no severe comorbidity. Chemotherapy: HD-methotrexate +/- doxorubicin +/- cisplatin +/- ifosfamide +/- BCD. Scheduled local therapy: Surgery. RESULTS: 925 evaluable patients from 101 institutions. Median age 15 years, m:f 1.4:1. Primary site: femur 510, tibia 251, humerus 100, fibula 51, other 13. Tumor-size < 1/3 of the involved bone 616, > or = 1/3 304. Definitive surgery in 903/925 cases, 443 limb salvage procedures. Good response (> 90% necrosis) in 469/806 (58.2%) evaluated tumors. Median follow-up for surviving patients: 5.42 years. Actuarial survival after 5 and 10 years: 72.5% (95%-CI 69.3-75.7) and 66.3% (62.5-70.0), relapse-free 62.1% (58.7-65.4) and 59.4% (55.8-63.0). 683/925 alive (601 first remission), 242 deceased (212 tumor progression, 30 other causes). 66.2% (97.3%) of all relapses within 2 (5) years. Prognosis correlates with tumor-size (< vs. > or = 1/3: 69.9% vs. 58.3% at 10 years) and -site (tibia: 74.2%, humerus: 54.5%) and -response (good vs. poor: 78.2% vs. 52.5%) (all p < 0.01). Actuarial 10-year survival by response grading I-VI according to Salzer-Kuntschik 80.9%, 82.8%, 71.1%, 60.7%, 47.7%, 27.3%. COSS-studies with preoperative 4-drug therapy more efficacious than less aggressive protocols. No impact of doxorubicin scheduling (sequential: rapid vs. 48 h-continuous infusion) or cisplatin scheduling (randomized: 5 h vs. 72 h-infusion) on prognosis detected. CONCLUSIONS: Intensive multiagent chemotherapy and delayed surgery for localized extremity osteosarcoma led to excellent oncologic results in the COSS-studies. Tumor-size, -site, and -response as well as the intensity of upfront chemotherapy correlated with outcome. Giving doxorubicin and cisplatin by continuous infusions did not result in discernible prognostic disadvantages.
Adolescent↗
Short-term culturing influences the number of bcr/abl-fused cells detected by fluorescence in situ hybridisation in bone marrow aspirates of CML patients.
Fluorescence in situ hybridisation (FISH) has been proven as a helpful tool in diagnosis and monitoring of bcr/abl fusion in chronic myelogeneous leukaemia (CML). Since long-term cultures are known to decrease the number of bcr/abl-fused cells, it is questionable whether similar effects are detectable in short-term cultures, a technique often preceding FISH analysis. Therefore, we evaluated bone marrow aspirates of 10 CML patients at biopsy and after culturing for between 24 and 144 h by FISH. The percentage of bcr/abl-fused cells in FISH varied between 15 and 70% at biopsy. In samples with 15 and 30% of aberrant cells at biopsy, an increase of about 20% per day was seen within the first 48 h. In longer lasting cultures, the percentage of leukaemic cells then asymptotically approached a value of 60-70%. In patients with 38 and 50% of bcr/abl-fused cells at biopsy, an increase of about 20% could be detected in the first 24 h. Then 65-70% of the cells already bore the bcr/abl fusion, and the percentage of leukaemic cells was almost constant for longer lasting cultures up to 144 h. In patients with a percentage of about 70% before culturing, no increase in positive cells was detected. These results emphasize the impact of short-term culturing on the number of bcr/abl-fused cells. In particular, the importance for monitoring CML patients is obvious. Therefore the effect described should be taken into account in order to avoid misinterpretation and incorrect therapy decisions in CML patients.
Adult↗
Microdissection of tissue sections: application to the molecular genetic characterisation of premalignant lesions.
The characterisation of the early molecular genetic events of tumor development depends on the selective procurement of histopathologically defined small cell populations from premalignant tissue. In order to obtain high-quality DNA, mRNA and proteins from these small tissue samples and even from single cells, tissue microdissection is one of the most useful techniques, becoming increasingly important for molecular pathologists. Using different microdissection techniques which allow the isolation of morphologically defined cell populations under direct visualisation, it is now feasible to study molecular genetic events that drive the multistep evolution in tumours. This review aims to present the current techniques of tissue microdissection and these techniques are discussed in the light of their ability to isolate premalignant cell populations in particular. Furthermore, we describe the subsequent application of several multiplex molecular analyses for characterising the microdissected premalignant cells. Applying these advanced techniques, alterations in the cellular DNA or the fluctuation of expressed genes that correlate with a particular stage of carcinogenesis can ultimately be compared within or between individual patients. Thus, these new technologies will have an enormous impact on molecular pathology with several diagnostic, prognostic and therapeutic implications.
Cell Separation↗
Detection of micrometastases in sentinel lymph nodes of the breast applying monoclonal antibodies AE1/AE3 to pancytokeratins.
Sentinel lymph node excision in breast cancer is a minimally invasive diagnostic procedure for accurate staging of the axilla and for avoiding unnecessary axillary dissection. In patients with palpable breast cancer we injected microcolloidal particles of human serum albumin labelled with technetium-99m the day before surgery. The sentinel node was detected intraoperatively with a handheld gammaprobe and then removed. Complete axillary dissection was performed and the nodes inspected by routine histological examination. The axillary lymph node status was correctly predicted by the sentinel node technique in 32 of 33 breast cancer patients. Two cases of micrometastases escaped routine histopathological detection but were identified by immunohistochemical analysis applying the antibody AE1/AE3 to pancytokeratins. Immunohistochemical examination of the sentinel node improves the diagnostic security of patients with breast carcinoma by detection of micrometastases.
Adult↗
Comparison of different histopathological methods for the examination of sentinel lymph nodes in breast cancer.
BACKGROUND: Sentinel lymphonodectomy is a new method for the classification of axillary lymph nodes in breast cancer. The optimum technique for the pathological examination of sentinel lymph nodes (SLNs) is still under debate. MATERIALS AND METHODS: Different histopathological techniques were evaluated in order to study their diagnostic accuracy regarding the detection of metastases in 49 SLNs of 40 breast cancer patients. RESULTS: In single hematoxylin and eosin (HE) stained paraffin sections 18 out of 40 patients showed positive SLNs and 8 out of 40 showed positive axillary lymph nodes (ALNs). Serial sections with a spacing of 150 microns between following sections revealed an additional 4 and 5 cases with positive SLNs and ALNs, respectively. Single HE frozen sections showed 11 tumor positive SLNs among 25 patients. The ultrarapid-immunohistochemistry on single frozen sections confirmed these data and detected one additional case with isolated tumor cells in the lymph node sinus. CONCLUSION: Histopathological nodal staging of SLNs should include serial sections with a spacing of 150 microns between sections as well as immunohistochemistry. The ultrarapid-immunohistochemistry is a sensitive method for the detection of minimal metastatic disease in SLNs and can be applied intraoperatively.
Adult↗
SV40 sequences in human osteosarcoma of German origin.
Simian virus 40 (SV40) is a dsDNA polyomavirus that induces osteosarcomas and mesotheliomas in hamsters and transforms many types of cells in tissue culture, including human cells. Osteosarcoma is a bone malignancy with multiple molecular lesions underlining progression from normal bone to osteosarcoma. Recent investigations have identified SV40 DNA sequences in osteosarcomas, suggesting that SV40 may contribute to tumor development. However, these studies also demonstrated that geographical differences exist between SV40 and tumor association. Our study analyzed 46 frozen German tumor specimens (42 osteosarcomas and 4 sarcomas initially suspected to be osteosarcomas) for the presence of SV40 DNA sequences by using PCR. Two different primer sets amplifying a 573 bp region of SV40 Tag gene with the complete intron sequence (SV.for 2/SV.rev) and a 172 bp region with no intron sequence (SV.5/SV.6) were used. DNA sequencing analysis verified the results. No SV40 sequences could be detected using the primer set SV.for 2/SV.rev, while 2 out of 42 osteosarcoma specimens and 1 out of 4 poorly-differentiated tumor specimens contained SV40 sequences, using the primer set SV.5/SV.6. From one of these two positive osteosarcomas, multiple tumor biopsies taken at different times during the dissection, including metastasis, tested positive for SV40. These results indicated that in Germany, only rare osteosarcomas can be linked to SV40. These results support previous findings involving geographical differences in the presence of SV40. Finally, the specific detection of SV40 sequences with multiple specimens from one of the two patients and the absence of SV40 sequences in all other samples underscores the specificity and reproducibility of this investigation and ruled out PCR contamination.
Adolescent↗