Gains and losses of DNA sequences in osteosarcomas by comparative genomic hybridization.

Our aim was to identify chromosomal regions that are likely to harbor previously unknown genes with an important role in the genesis of osteosarcoma. Comparative genomic hybridization was used to screen for losses and gains of DNA sequences along all chromosome arms in 11 tumors. Extensive genetic aberrations, with an average of 11 changes/tumor (range, 1-20), were found in 10 of the 11 specimens. High level amplifications of small chromosomal regions were detected in eight tumors. These involved the 12q12-q13 region (known to contain the SAS-MDM2 locus) and several previously unreported amplification sites such as 17p11-p12, 3q26, and Xq12. When all DNA sequence gains were evaluated, the gains at 8q and Xp were most common (45%). The most common losses of DNA sequences were seen at 2q, 6q, 8p, and 10p (36%). In conclusion, despite the very complex pattern of genetic changes in osteosarcomas, certain chromosomal regions appear to be affected more often than others. Most of these regions have not previously been reported to be implicated in osteosarcomas and may thus highlight locations of novel genes with an important role in the development and progression of these tumors.

Surgical treatment of myeloma of bone.

33 patients treated operatively for plasma cell disease were analysed. There were 21 men and 12 women with an average age of 54 years. There was an undefined bone tumour in 23 cases, and a pathologic fracture in 10 cases. In only 6 cases was the diagnosis known before the operation. The primary tumour localisations were: vertebral column in 13, pelvis in 7, femur in 6, humerus in 2, rib in 1, tibia in 1, fibula in 1, scapula in 1 and olecranon in 1 case. 16 diagnostic biopsies were taken. Vertebral tumours were mainly evacuated or decompressed, combined with a stabilising procedure in 8 cases. A total of six endoprostheses, five to the femur and one to the humerus were performed. Two primarily wide resections, to the fibula and to the scapula were done. There were no locally recurring tumours during a mean follow-up time of 4 years and 2 months, and we conclude that operative and oncologic treatment is successful in providing the patient with a stable, pain-free locomotive system.

Chordoma with a massive spindle-cell sarcomatous transformation. A light- and electron-microscopic and immunohistological study.

A 26-year-old woman was operated on for a bulky tumor in the sacral region; she died of massive local tumor recurrence and pulmonary metastases 3 months later. Most of the original tumor showed a highly cellular spindle-cell sarcoma compatible with a fibrosarcoma of a high grade of malignancy. In a few small areas of the tumor, a chordoma-like pattern surrounded by growth of spindle-cell sarcoma was found. The spindle-cell component exhibited vimentin positivity in all tumor cells, but many cells were also cytokeratin-positive. The chordoma-like areas showed cytokeratin in all tumor cells. The chordoma-like areas, but not the spindle-cell areas also were positive for epithelial membrane antigen and S-100 protein. This case indicates that the sarcomatous change associated with chordoma may contain keratins as a sign of epithelial differentiation, and may thus represent sarcomatous transformation of chordoma cells, rather than a coincidental soft-tissue sarcoma or collision tumor.

Clinical significance of genetic imbalances revealed by comparative genomic hybridization in chondrosarcomas.

DNA copy number changes were studied by comparative genomic hybridization (CGH) in 50 chondrosarcoma samples from 45 patients. Mean number of genetic aberrations in primary tumors was 4.8 +/- 1.8. The most frequently gained regions were 20q12-qter (37%), 20q (32%), 8q24.1-qter (27%), 20p (24%), and 14q24-qter (24%). Losses were 5.5 times less frequent than gains and observed mainly at Xcen-q21, 6cen-q22, and 18cen-q11.2 (11% each). Recurrent and metastatic tumors showed a mean of 4.0 +/- 2.2 aberrations per sample. The most frequently gained regions were chromosome 7 (4 cases), 5q14-q32 (4 cases), 6p (3 cases), and 12q (3 cases). Losses of DNA sequences were 3.4 times less frequent than gains. Histological tumor grade was significantly associated with metastasis-free survival (P = .002) and overall survival (P = .003), being the strongest prognostic factor tested. A statistically significant correlation was found between gain at 8q24.1-qter and shorter overall survival (P = .01) but not with local recurrence or metastasis-free survival. Gain at 14q24-qter was associated with a trend to shorter overall survival (P = .05) but neither with an increased risk for local recurrence nor with metastasis-free survival. In a multivariate analysis, only the tumor grade associated with overall survival (P = .02). In a multivariate analysis together with the tumor grade, gain at 8q24.1-qter did not retain its significance (P = .44), indicating that this imbalance is not an independent prognostic factor.

A recurrent chondromyxoid fibroma with chromosome aberrations ins(5;2)(q13;p21p25) and 2p deletion: a case report.

We report a patient with a recurrent chondromyxoid fibroma, a rare benign tumor of the bone with clonal aberrations in chromosomes 2 and 5. Karyotyping, chromosome painting, interphase cytogenetics by in situ hybridization, and DNA flow cytometry were used. The karyotype was interpreted as 46,XX,der(2)ins(5;2)(q13;p21p25),der(2)ins(5;2)(q13;p21p25), der(5)ins(5;2) (q13;p21p25).

Cytogenetic study of 249 consecutive patients examined for a bone tumor.

Chromosome analysis was performed on 304 samples of 249 consecutive patients examined for a possible bone tumor. The series consisted of 86 nonneoplastic disorders, 108 benign and 78 malignant primary bone tumors, and 32 other bone malignancies. In the group of nonneoplastic disorders, one sample from an infectious lesion demonstrated a clonal chromosome aberration, i.e., additional material in the short arm of chromosome 1. Simple clonal aberrations were noted in six of 75 successfully cultured benign tumors, e.g., a chondromyxoid fibroma with an insertion type translocation from 2p21p25 to 5q13 and 2p deletion and a nonossifying fibroma with del(4)(p14). Complex clonal aberrations were evident in 21 of 54 successfully cultured malignant primary bone tumors and eight of 21 secondary bone malignancies. The complexity of clonal aberrations correlated with the grade of malignancy as the osteosarcomas and chondrosarcomas of high-grade demonstrated chaotic abnormalities. Six Ewing's sarcomas demonstrated the t(11;22)(q24;q12); in one this was the sole abnormality, and in five additional changes were evident: der(1;16)(q10;p10) in one. Homogeneously staining elongated areas interpreted as HSR were observed in three patients, all of whom had a highly malignant tumor. The most frequent nonclonal abnormality was telomeric association, which was observed mainly in giant cell tumors.

No DNA copy number changes in osteochondromas: a comparative genomic hybridization study.

Cytogenetic changes in osteochondroma samples were studied by comparative genomic hybridization and by chromosome banding. No DNA copy number changes (15 patients) or chromosomal aberrations (9 patients) were observed in any of the patients.

Treatment of solitary plasmacytoma of the femur. A case report.

In radical tumour surgery the inevitable bone loss is considerable. If in addition the local tissues at the tumour site have been damaged by radiation therapy, massive endoprosthesis or even amputation should be considered. In cases with large bone loss, a vascularized fibular graft is one possibility to bridge the resected region. A case of a young man with a solitary plasmacytoma of the upper femoral diaphysis is presented. The lesion was reoperated 3 years after the primary diagnosis with resection of the non-ossified tumour site and reconstruction with a vascularized fibular graft.

Giant cell tumours of bone. A DNA-flow cytometric study.

The variable clinical course and locally aggressive growth of giant cell tumours of bone cause problems in planning treatment. Curettage and bone grafting is the commonest form of surgical treatment, but recurrence rates as high as 40 per cent have been reported. Thirty-four patients with a GCT were treated in our clinic between years 1982 and 1990. There were three gr 1, 24 gr II and 7 grade III tumours. Operative treatment was carried out in 31. DNA flow cytometry was performed from the sample in 28 patients. The type of treatment played an important role in tumour prognosis: there were 10 recurrences after 18 intralesional procedures, but only one recurrence after tumour resection. DNA flow cytometry was not helpful in predicting tumour behaviour.

Malignant fibrous histiocytoma arising in a previous site of fracture and osteomyelitis.

Malignant fibrous histiocytomas (MFH) are known to arise in benign pre-existing conditions. We present here a case where MFH occurred more than 40 years after a fracture and infection. The diagnosis was established only after repeated clinical and histopathologic examinations. DNA flow cytometric and chromosomal analyses were also done.

Chondrosarcoma of bone. A clinical and DNA flow cytometric study.

Fifty-two patients with chondrosarcoma were treated in our hospital between 1981 and 1992. Tumours from 21 of the patients were histologically grade I, 19 were grade II, 10 were grade III and two were grade IV. Forty-seven patients were treated surgically and samples for flow cytometric DNA analysis were taken from 38 tumours with a successful measurement in 37 cases. The mean follow-up time was 40 months and during this period 17 of the patients treated surgically developed a local recurrence or metastasis. In one case, a highly anaplastic sarcoma developed after radiation treatment. Only two of the 19 diploid tumours recurred, whereas eight of the 18 aneuploid tumours showed a recurrence. These results indicate that DNA flow cytometry is of prognostic significance in chondrosarcoma and may aid in planning the treatment.

Ten years' experience in patients with osteogenic sarcoma in Finland.

Ten years' results of 56 patients with high grade osteogenic sarcoma are reported. Fifty-two patients had M0 disease. Immediately after open biopsy the patients were treated with chemotherapy using modified Rosen's protocols T4, T7 and T10. The primary tumor was adequately removed in most patients. Six children were treated with limb saving. The actuarial and disease-free survival was 80% after 1 year, and 73% to 8 years. Two patients died because of toxic side effects of chemotherapy, one of septicemia, the other of late cardiac failure secondary to doxorubicin.

Relationship of the Pain Disability Index (PDI) and the Oswestry Disability Questionnaire (ODQ) with three dynamic physical tests in a group of patients with chronic low-back and leg pain.

OBJECTIVE: To determine the relationship between overall disability in daily activities, assessed with the Pain Disability Index (PDI) and the Oswestry Disability Questionnaire (ODQ), and impaired performance on three physical tests in patients with chronic low-back pain. DESIGN AND SUBJECTS: The PDI and ODQ were administered in a cross-sectional study, before beginning a back rehabilitation program, to 45 patients with low-back pain of > or = 3 months' duration, with or without radiation to the legs. All patients also performed repetitive sit-up, arch-up, and squatting tests. SETTING: Tertiary care center. RESULTS: Modestly significant (p < 0.05) or significant (p < 0.01) inverse correlations (Pearson's r = 0.30-0.41) were noted between the PDI and the ODQ and all three physical performance tests. When normative data were used, the correlation (Spearman's rs = -0.45) between PDI and the squatting test remained significant (p < 0.01), whereas it was modestly significant (rs = -0.33, p < 0.05) between the ODQ and squatting test and between the PDI and arch-up test (rs = -0.35, p < 0.05). Compared with patients presently working, those on sick leave had significantly higher scores on the PDI and ODQ (Wilcoxon's two-sample test: p < 0.001) and also significantly worse performance on all physical tests (p < 0.001). CONCLUSIONS: The PDI and ODQ, as measures of self-perceived disability, and impaired performance on repetitive squatting, arch-up, and sit-up tests, as measures of physical capability, show some overlap in low-back-pain patients. Both types of disability measures are clearly influenced by the patient's work status.

Intercorrelation and test-retest reliability of the Pain Disability Index (PDI) and the Oswestry Disability Questionnaire (ODQ) and their correlation with pain intensity in low back pain patients.

OBJECTIVE: To determine the intercorrelation between subjective disability, as assessed with the Pain Disability Index (PDI) and the Oswestry Disability Questionnaire (ODQ) and their correlation with visual analogue scale (VAS) pain intensity ratings. DESIGN AND SUBJECTS: Questionnaires were administered to 94 patients with chronic low back pain with or without radiation into the legs of at least 3 months' duration. SETTING: Tertiary care center. RESULTS: High correlations were noted between the ODQ and PDI (r = 0.83) and PDI factor 1 (r = 0.84), a subscale of the PDI. Lower correlations were noted between pain intensity (VAS) scores and the ODQ (r = 0.62) and the PDI (r = 0.69). A weaker correlation (r = 0.41) was noted between the ODQ and PDI factor 2. Intraclass correlation coefficients (ICC) for test-retest reliability in 20 patients (time interval 1 week) were for the ODQ ICC = 0.83, PDI ICC = 0.91, PDI percentage score ICC = 0.91, PDI factor 1 ICC = 0.87, and PDI factor 2 ICC = 0.73, respectively. CONCLUSIONS: The present results suggest that either the PDI or the percentage score PDI and also the even shorter-to-administer PDI factor 1 may be useful and reliable tests for the assessment of subjective disability in low back pain patients. As noted by the moderate intercorrelations with pain intensity scores, both the PDI and the ODQ address a broader concept of disability than that directly related to pain intensity.

Structural changes in intact tubular bone after application of rigid plates with and without compression.

The influence of a rigid plate on the structure of intact tubular bone was studied in forty adult rabbits using a four-hole commercial-steel DCP-ASIF plate attached to each tibiofibular bone, with and without compression. The morphological changes in the bone underlying the plate were examined and were assessed quantitatively at one day to thirty-six weeks after operation. The rigid plate induced porotic changes in the cortical bone, which were evident three weeks after the operation and increased rapidly between twelve and eighteen weeks. At thirty-six weeks, resorption cavities occupied 40.6 per cent of the cortical wall of the bones plated with compression and 41.2 per cent of the wall of those plated without compression. The changes were visible slightly earlier in the presence of compression. A rigid plate brought about a statistically significant increase both in the outer diameter of the bone and in the diameter of the medullary space as a result of subperiosteal new-bone formation and concomitant subendosteal bone resorption.

Immunohistochemical demonstration of nociceptors in the ligamentous structures of the lumbar spine.

Substance P, a physiologically potent neuropeptide is known to participate in the sensory, and especially nociceptive, transmission of neural impulses. On histologic grounds, the nerve terminals of the sinuvertebral nerve formerly have been suggested to be sensory in character and to mediate the low-back pain syndrome. Samples of paramedullary ligamentous structures were collected on disc operations. A positive immunoreaction as an indicator of substance P was confirmed in some nerve terminals of the posterior longitudinal ligament. Neither the yellow ligament nor the intervertebral disc showed such nociceptive-type nerves.

Inflammatory cells, motor weakness, and straight leg raising in transligamentous disc herniations.

STUDY DESIGN: Possible statistically significant relationships between inflammatory cells and either motor weakness or straight leg raising were determined. OBJECTIVES: To look for any clinically relevant links between inflammatory cells in disc herniations and signs of radiculopathy. SUMMARY OF BACKGROUND DATA: Many studies have during recent years shown a presence of various types of inflammatory cells in disc herniations, but their clinical relevance has been questioned. To be clinically relevant, a presence of inflammatory cells should show a clear relationship to clinical evidence of nerve root involvement. Macrophages repeatedly demonstrated in a high proportion of disc herniations studied are of particular interest. Their major role may be in disc herniations tissue resorption and not in sciatica. METHODS: A total of 96 disc herniations, all transligamentous, were analyzed by immunohistochemistry for presence of macrophages, T or B lymphocytes, and activated T lymphocytes separately. From recorded patient data, motor weakness and straight leg raising data were compared with a presence or absence of abundant (+ = at least 20 cells in a group) inflammatory cells. When not abundant, inflammatory cells were classified as "only few cells" (+) and grouped together with "no cells" (-). Patients with or without motor weakness were compared. Straight leg raising was compared for a positive (at <70 degrees ) or a negative test, and separately using the median as cut-off value. Groups were compared by chi-square analysis with the level of statistical significance set at P<0.05. RESULTS: None of the four inflammatory cell types showed any significant association with motor weakness. Nor was any association observed when comparing positive and negative straight leg raising. With the median (straight leg raising = 47.5 degrees ) as cut-off, only activated T cells showed a weak (chi2 = 4.40, P<0.05) relationship with tighter straight leg raising, but none of the other cell types did. Even when straight leg raising was < 47.5 degrees, three times more disc herniations lacked (n = 34) inflammatory cells than showed (n = 13) inflammation. In a subgroup of only sequestrated discs, the findings were similar. However, in the patients with a bilaterally positive straight leg raising (n = 25), the prevalence of at least one inflammatory cell type was much higher in sequestrated discs (80%) than in extrusions (33%). This may suggest more subtle interrelationships between type of disc herniation, straight leg raising, and inflammatory cells. CONCLUSIONS: The results of this study do not support a clinically relevant role for disc herniation inflammatory cells in sciatica. For the cells to be clinically relevant, a strong relationship between a presence of inflammatory cells and either or both of motor weakness and a tight straight leg raising should have been observed. The authors conclude that macrophages, which have been demonstrated in a high proportion of disc herniations in previous studies, are probably more important for disc tissue resorption processes than for producing sciatica. Other types of inflammatory cells are more rarely observed and may have no clinical meaning at all. However, more subtle interrelationships, considering the various types of disc herniations, should be further explored.

The role of mast cells in disc herniation inflammation.

STUDY DESIGN: A study of herniated lumbar disc tissue samples and control disc material to determine the presence of mast cells in disc herniations. OBJECTIVES: To analyze whether mast cells have any involvement in disc herniation pathophysiology and lumbar pain, because mast cells may have an important role in acute and chronic inflammatory responses. SUMMARY OF BACKGROUND DATA: Studies of inflammatory cells, biochemical mediators of inflammation, and tissue degrading enzymes have suggested that these factors may be involved--and perhaps play an important role--in the pathophysiology of lumbar pain and radiculopathy. Mast cells are known to play an important role in acute and chronic inflammatory responses. It was therefore of interest to clarify their possible role in intervertebral disc herniation inflammation. METHODS: Fifty herniated lumbar discs from 50 patients who had undergone disc surgery and three normal control discs were obtained. Sections from every disc then were examined histologically and immunocytochemically for mast cells by using monoclonal antibodies to either of two types of specific proteases of mast cells, tryptase and chymase. RESULTS: By none of the methods could any mast cells be observed in any of the control disc samples. With toluidine blue staining, mast cells were observed in 9 of 50 (18%) of discs. Mast cells immunoreactive to either tryptase or chymase were observed in 10 of 50 disc samples (20%) and immunoreactive for tryptase and chymase simultaneously in 4 of 50 disc samples (8%). However, the majority of the samples studied (80%) demonstrated immunoreactivity to neither tryptase nor chymase. Among the samples studied were five disc protrusions that totally lacked mast cells. CONCLUSIONS: A minority of disc herniations exhibited mast cells, as verified by toluidine blue staining and immunocytochemistry. The results may suggest a role of mast cells in intervertebral disc herniation inflammation, but only in a subset of these cases. Massive infiltration by mast cells never was observed.

A comparative immunohistochemical study of inflammatory cells in acute-stage and chronic-stage disc herniations.

STUDY DESIGN: Herniated lumbar disc specimens were obtained from patients undergoing surgical discectomy for persistent radicular pain (radiculopathy) and stained for inflammatory cells to determine their occurrence in relation to the duration of radicular pain and to analyze the role of the time factor in the inflammatory response. OBJECTIVES: To analyze the presence of inflammatory cells and their involvement in the pathophysiology of radicular pain and to determine whether there is a clear difference in the occurrence of inflammatory cells between the earlier phase of radicular pain (after herniation) and the later chronic stage. SUMMARY OF BACKGROUND DATA: Previously, inflammatory cells were reported in herniated disc tissues, and macrophages were most prevalent. Biologically active inflammatory mediators have also been repeatedly observed. However, there have been no observations regarding possible differences in the occurrence of inflammatory cells in radicular pain of different durations. METHODS: Forty-four herniated lumbar discs were obtained from 44 patients undergoing disc surgery. Two groups of 22 age- and gender-matched patients with comparable affected disc levels were studied. In the first group (acute group) pain duration ranged from 3 days to 21 days. In the second group (chronic group) pain duration was 6 months or longer. All disc herniation specimens were subjected to indirect immunocytochemistry to study and compare the presence of inflammatory cells. RESULTS: Inflammatory cells, predominantly macrophages, were observed in both groups. Macrophages were abundantly present in eight (36%) disc samples in the acute group; in three (14%) samples only few scattered macrophages were observed. In the chronic group, in nine (41%) disc samples, abundant macrophages were observed; in six (27%) there were a few scattered macrophages. In the acute group, in three (14%) disc samples abundant activated T lymphocytes were observed; in two (9%) there were only a few activated T lymphocytes, whereas in the chronic group abundant activated T lymphocytes were not seen; only a few scattered activated T lymphocytes were observed in five (23%) disc tissue samples. In two (9%) samples in the acute group, B cells were abundantly present, and in two (9%) only a few B cells were observed. In the chronic group, abundant B cells were seen in no samples, and only a few B cells were noted in one (5%) sample. Only in the acute group and only in lateral disc herniations were abundant lymphocytes observed. In disc samples from intraspinal herniations, acute and chronic, there were only abundant macrophages, not lymphocytes. CONCLUSIONS: Because of the small size of the study groups and the low prevalence particularly of lymphocytes in both groups, no major group differences were noted. The prevalence of macrophages was highest, similar in both groups, and was similar to the results in prior studies. The results indicate no major differences in the occurrence of inflammatory cells in acute and chronic disc herniations. They also indicate that only macrophages may have a clinical relevance in disc tissue inflammation.

An immunohistochemical study of nerve structures in the anulus fibrosus of human normal lumbar intervertebral discs.

STUDY DESIGN: The innervation of the anulus fibrosus of human macroscopically normal intervertebral discs from five patients was investigated immunohistochemically. OBJECTIVES: Immunoreactivity to general nerve markers (synaptophysin and protein gene product 9.5) and to neuropeptides (substance P and C-flanking peptide of neuropeptide Y) was studied. SUMMARY OF BACKGROUND DATA: In the lumbar disc of a newborn, free nerve endings have been demonstrated in the outer layers of anulus fibrosus. In degenerated and herniated discs, nerve structures have been shown to penetrate deeper into the anulus fibrosus. There are only a few studies on the innervation of normal adult intervertebral disc tissue. METHODS: Thin frozen sections of human normal lumbar intervertebral disc tissue were immunostained for general nerve markers and neuropeptides. RESULTS: Synaptophysin and protein gene product 9.5 immunoreactive nerve structures were observed penetrating 3.5 mm and 1.1 mm into the anulus, respectively. Immunoreactivity to C-flanking peptide of neuropeptide Y and substance P were observed at a maximum depth of 0.9 and 0.5 mm in the anulus, respectively. Antibodies to the former have been used to study sympathetic nerves, whereas substance P is a transmitter present in sensory nerves. CONCLUSIONS: In anulus fibrosus samples from macroscopically normal discs, a general marker for nerve endings can be found at a depth of a few millimeters, whereas neuropeptide markers show nerves only in the outermost layers of the anulus fibrosus. This absence of demonstrable nerves in deeper anulus fibrosus in normal discs is probably not a methodologic artifact, because blood vessels have also been demonstrated only at the disc surface. It is, however, possible that neuropeptide nerves also penetrate to a depth of a few millimeters, but that methodologic limitations permit the visualization of only the neuropeptide nerves closest to the disc surface. The results of the present study lend support to previous suggestions that, except at the surface, a normal intervertebral disc is almost without innervation.